Brine shrimp lethality test active constituents and new highly oxygenated seco-prezizaane-type sesquiterpenes from Illicium merrillianum

In the study of bioactive substances in Illicium plants, the methanol extract of I. merrillianum showed brine shrimp lethality test (BST) activity at 200 microg/ml. Bioassay-guided fractionation of the BST active fractions resulted in the isolation of 4-O-methyleudesm-11-en-4alpha-ol, eudesmol-11-en-4alpha-ol and (-)-hinokinin as potent BST active compounds. On the other hand, four new highly oxygenated seco-prezizaane-type sesquiterpenes, merrilliortholactone (1), 2alpha-hydroxycycloparvifloralone (2), 2alpha-hydroxycycloparviflorolide (3), and 2alpha-hydroxyanisatin (4) were isolated from the BST-inactive polar fractions. The structures of new compounds were elucidated by extensive analyses of spectral data. Furthermore, the absolute configuration of 3 was established by the modified Mosher's method. Compounds 1--4 showed neither BST activity at 100 microg/ml nor neurite outgrowth-promoting activity.     

A new fatty acid ester of triterpenoid from Celastrus rosthornianus with anti-tumor activitives

The methanol extract of the stalks of Celastrus rosthornianus Loes. afforded a new fatty acid ester of triterpene: 1beta,3beta-dihydroxy-olean-9(11),12-dienyl-3-palmitate (1), and three known compounds beta-amyrin palmitate (2), 3beta-hydroxy-11-oxo-olean-12-enyl-3-palmitate (3) and lupeol palmitate (4). Their structures were established by HRMS, 1D and 2D NMR experiments, as well as, by chemical degradation. The new compound showed cytotoxicity against human cervical squamous carcinoma (Hela) cells.     

Structures of 1,4-benzodioxane derivatives from the seeds of Phytolacca americana and their neuritogenic activity in primary cultured rat cortical neurons

The methanol extract of the seeds of Phytolacca americana was reinvestigated to yield three new 1,4-benzodioxane-type compounds, americanoic acid methyl ester (1), isoamericanoic acid A methyl ester (2), and 9'-O-methylamericanol A (3) along with the previously isolated neolignans 6-9. The structures of 1-3 were characterized by 2D NMR and long-range selective proton-decoupling (LSPD) techniques. The neuritogenic effects of compounds 1-3, and dicarboxilic acids 4 and 5, which had been previously synthesized with horseradish peroxidase-catalyzed oxidative coupling of caffeic acid, were examined in primary cultured rat cortical neurons. Americanoic acid A methyl ester (1) exhibited neurite outgrowth-promoting activity at concentration of 0.01-1.0 microM, whereas dicarboxilic acids 4 and 5 were found to induce neuritogenesis dose dependently at the concentrations from 0.1 microM to 10 microM.     

Isolation and structure of a monomethylated ganglioside possessing neuritogenic activity from the ovary of the sea urchin Diadema setosum

A new monomethylated ganglioside, DSG-A (3), was obtained, together with four known gangliosides, compounds (1, 2, 4, 5), from the lipid fraction of the chloroform/methanol extract of the ovary of the sea urchin Diadema setosum. The structures of the new ganglioside was determined on the basis of chemical and spectroscopic evidence to be 1-O-[9-O-methyl-(N-acetyl-alpha-D-neuraminosyl)-(2-->6)-beta-D-glucopyranosyl]-ceramide (3). The ceramide moiety of 3 was composed of C18-phytosphingosine base, and 2-hydroxy and nonhydroxylated fatty acid units. These gangliosides showed neuritogenic activity toward the rat pheochromocytoma cell line PC-12 in the presence of nerve growth factor, in which compound 3 showed the most potent activity.     

New 3',8'-linked biflavonoids from Selaginella uncinata displaying protective effect against anoxia

Seven 3',8'-linked bioflavonoids, including one new compound, (2'S)-2', 3'-dihydroamentoflavone-4'-methyl ether and six known compounds: (2S)-2,3- dihydroamentoflavone-4'-methyl ether, (2S,2'S)-2,3,2',3'-tetrahydroamento- flavone-4'-methyl ether, (2S,2'S)-tetrahydroamentoflavone, (2S)-2,3-dihydro- amentoflavone and (2'S)-2',3'-dihydroamentoflavone (6) and amentoflavone, were isolated from the 60% ethanolic extract of Selaginella uncinata (Desv.) Spring. The structures of these compounds were elucidated mainly by analysis of their 1D and 2D NMR spectroscopic data, and their absolute configurations were determined by circular-dichroism (CD) spectroscopy. All the seven compounds showed protective effect against anoxia in the anoxic PC12 cells assay, in which compound 6 displayed particularly potent activity.     

Chemical constituents and antioxidant activity of Ficus callosa

One new megastigmane glycoside, ficalloside (1), and eleven known compounds, were isolated from methanol extract of Ficus callosa leaves by repeated column chromatography. Their structures were established on the basis of spectral and chemical evidence. The antioxidant activities of these compounds were measured using the oxygen radical absorbance capacity (ORAC) assay. Compound 8 exhibited potent antioxidant activity of 10.6 microM trolox equivalents at the concentration of 2 microM. At this concentration, compounds 4-7 and 9-12 showed significant antioxidant activity with ranging of 2.1-6.1 microM trolox equivalents.     

Iridoids and sesquiterpenoids with NGF-potentiating activity from the rhizomes and roots of Valeriana fauriei

A new iridoid glycoside, 10-isovaleryl kanokoside C (1), and a new sesquiterpene (2) together with two known compounds (3, 4) were isolated from the rhizomes and roots of Valeriana fauriei. Their structures were elucidated on the basis of spectroscopic analysis. Compounds 2 and 4 showed enhancing activity of nerve growth factor (NGF)-induced neurite outgrowth in PC 12D cells.     

Beyerane derivatives and a sesquiterpene dimer from Japanese Cypress (Chamaecyparis obtusa)

In the course of studies on biological active constituents from woody plants, we previously reported the isolation of many lignan derivatives as neurite outgrowth-promoting compounds from an ethyl acetate soluble fraction of Japanese Cypress (Chamaecyparis obtusa). Further chemical investigation on the residual parts of the ethyl acetate soluble fraction of a methanol extract of Japanese Cypress resulted in the isolation of four new beyerene type derivatives and a novel sesquiterpene dimer formed between cryptomeridiol and hinokiic acid. Their structures were elucidated as 18-O-(Z)-p-coumaroylbeyer-15-ene-18-ol (1), 18-O-(E)-p-coumaroylbeyer-15-ene-18-ol (2), 18-O-(E)-p-coumaroylbeyer-15-ene-11beta,18-diol (3), 18-O-(Z)-p-coumaroylbeyer-15-ene-11beta,18-diol (4) and ent-cryptomeridiol-4-yl-hinokiiate (5) by (1)H-NMR, (13)C-NMR, 2D-NMR, and HR-MS spectral analysis.     

Degraded and oxetane-bearing limonoids from the roots of Melia azedarach

Brine shrimp lethality test (BST)-guided fractionation of a methanol extract of the roots of Melia azedarach resulted in the isolation of two new limonoids, 9alpha-hydroxy-12alpha-acetoxyfraxinellone (1) and 7,14-epoxy-azedarachin B (2), together with the known compounds, 12alpha-hydroxyfraxinellone (4), 9alpha-hydroxyfraxinellone (5), azedarachin B (6), and neoazedarachin B (7). The structures of 1 and 2 were elucidated by analysis of spectroscopic data and comparison of their NMR data with those of the known compounds. Compounds 1, 2 and 4-7 exhibited significant activity in the BST, in particular, azedarachin B (6) showed remarkable BST activity with an LC(50) value of 0.0098 microM.     

In vitro leishmanicidal constituents of Millettia pendula

The in vitro leishmanicidal constituents of Millettia pendula were examined. Two new compounds, 1 (millettilone A) and 2 (millettilone B), were isolated from the methanol extract of M. pendula, together with six known compounds: 3R-claussequinone (3), pendulone (4), secundiflorol I (5), 3,8-dihydroxy-9-methoxypterocarpan (6), 3,10-dihydroxy-7,9-dimethoxypterocarpan (7), and formononetin (8). Among these, pendulone showed the most potent leishmanicidal activity. Compound 2 was found to be a purple pigment in this heartwood. Their chemical structures were elucidated using spectral methods.     

A new monoterpenoid glycoside from Myrica esculenta and the inhibition of angiotensin I-converting enzyme

One new monoterpenoid glycoside, myresculoside (1), and eleven known compounds, were isolated from methanol extract of Myrica esculenta leaves by repeated column chromatography. The effects of these compounds on angiotensin I-converting enzyme (ACE) inhibition were investigated. Compounds 3 and 4 showed the most potent ACE inhibition with rates of 29.97% and 25.63% at concentration of 100 μM, respectively. Compounds 5, 6, and 11 showed weak activity with inhibitory rates of 0.07-1.41% at concentration of 100 μM.     

Constituents of Holothuroidea, 18. Isolation and structure of biologically active disialo- and trisialo-gangliosides from the sea cucumber Cucumaria echinata

Three new disialo- and trisialo-gangliosides, CEG-6 (6), CEG-8 (8), and CEG-9 (9), were obtained, together with one known ganglioside, HLG-3 (7), from the lipid fraction of the chloroform/methanol extract of the sea cucumber Cucumaria echinata. The structures of the new gangliosides were determined on the basis of chemical and spectroscopic evidence to be 1-O-[alpha-L-fucopyranosyl-(1-->11)-(N-glycolyl-alpha-D-neuraminosyl)-(2-->4)-(N-acetyl-alpha-D-neuraminosyl)-(2-->6)-beta-D-glucopyranosyl]-ceramide (6) and 1-O-[(N-glycolyl-D-neuraminosyl)-(2-->11)-(N-glycolyl-D-neuraminosyl)-(2-->4)-(N-acetyl-D-neuraminosyl)-(2-->6)-D-glucopyranosyl]-ceramide (8, 9). The ceramide moieties of each compound were composed of an homogeneous sphingosine or phytosphingosine base and heterogeneous 2-hydroxy or nonhydroxylated fatty acid units. These gangliosides showed neuritogenic activity toward the rat pheochromocytoma cell line PC-12 in the presence of nerve growth factor.     

Clerodane diterpenoids and flavonoids with NGF-potentiating activity from the aerial parts of Baccharis gaudichaudiana

A new clerodane diterpene, 15-hydroxy-16-acetoxy-ent-clerod-3-en-18-oic acid (1), together with three known clerodane diterpenes (2-4) and three known flavones (5-7), were isolated from the aerial parts of Baccharis gaudichaudiana. Their structures were elucidated on the basis of spectroscopic analysis. Compounds 2, 3, and 5 showed enhancing activity of nerve growth factor (NGF)-induced neurite outgrowth in PC 12D cells.     

Sesquiterpenoids of Illicium jiadifengpi and Their Effects on NGF‐induced Neurite Outgrowth in PC12 Cells

Two new compounds, jiadifenlactone acid monomethyl ester ( 1 ) and jiadifenin ( 3 ), and five known compounds were isolated from the fruits of I. jiadifengpi . Their structures were determined using spectroscopic techniques, including 1D, 2D NMR and HR‐ESI‐MS experiments. The relative stereochemistry of jiadifenlactone acid monometheyl ester ( 1 ) was further confirmed by X‐ray crystallographic data. All isolates were evaluated for their effects on nerve growth factor (NGF)‐mediated neurite outgrowth in pheochromocytoma (PC12) cells and two compounds showed promoting effects.     

Nitric oxide (NO) production inhibitory constituents of Tabebuia avellanedae from Brazil

From the water extract of Brazilian Tabebuia avellanedae, two new iridoids (1, 2) and a new phenylethanoid glycoside (3) have been isolated together with twelve known compounds (4-15). Their structures were determined based on the spectroscopic data. The isolated compounds inhibited nitric oxide (NO) production in lipopolysaccharide (LPS)-activated macrophage-like J774.1 cells. Compounds 1, 3, 10, 11, and 12 showed inhibitory activities more potent (IC50, 13.8-26.1 microg/ml) than a positive control N(G)-monomethyl-L-arginine (L-NMMA; IC50, 27.4 microg/ml).     

New diarylheptanoids from Alnus japonica and their antioxidative activity

In the course of research on the bioactive constituents of woody plants in the Cyugoku area of Japan, a methanol extract of the leaves of Alnus japonica were found to have strong antioxidative activity. Ethyl acetate soluble and n-buthanol soluble fractions of the methanol extract had a potent antioxidative effect. Both fractions were purified by silica gel column chromatography and HPLC using an ODS column to give four new diarylheptanoids along with known diarylheptanoids and flavonoids. These new compounds were elucidated to be 7-(3,4-dihydroxyphenyl)-5-hydroxy-1-(4-hydroxyphenyl)-3-heptanone-5-O-beta-D-xylopyranoside (1), 1-(3,4-dihydroxyphenyl)-5-hydroxy-7-(4-hydroxyphenyl)-3-heptanone-5-O-beta-D-xylopyranoside (2), 1,7-bis-(3,4-dihydroxyphenyl)-5-hydroxy-3-heptanone-5-O-[2-(2-methylbutenoyl)]-beta-D-xylopyranoside (3) and 1,7-bis-(3,4-dihydroxyphenyl)-5-methoxy-3-heptanone (4) using spectral methods and especially 1H-, 13C-NMR and 2D-NMR measurements. The isolated compounds including their main constituent, oregonin (5), were tested for antioxidative activity. Some of these compounds having two catechol structures showed potent antioxidative activity. Compounds having one catechol structure showed moderate antioxidative activity, but a peracetate of 5 having no catechol structure exhibited no antioxidative activity. Thus the catechol structure of the diarylheptanoids is indispensable for antioxidative activity.     

Anti-plasmodial and cholinesterase inhibiting activities of some constituents of Psorospermum glaberrimum

Glaberianthrone (1), a new bianthrone was isolated from the hexane extract of the stem bark of Psorospermum glaberrimum together with thirteen known compounds: 3-geranyloxyemodin anthrone (2), friedelan-3-one (3), 3-prenyloxyemodin anthrone (4), 3-geranyloxyemodin (5), 3-prenyloxyemodin (6), friedelan-3-ol (7), acetylvismione D (8), betulinic acid (9), 2-geranylemodin (10), bianthrone A2b (11), bianthrone 1a (12), emodin (13) and 2-prenylemodin (14). The structures of the isolated compounds were established by means of spectroscopic methods. The extracts and the isolated compounds were tested in vitro for their anti-plasmodial activity against Plasmodium falciparum (chloroquine resistant strain W2) and for their acetyl- and butyrylcholinesterase inhibitory properties. The n-hexane extract showed good anti-plasmodial activity against P. falciparum W2 strain, with IC(50) of 0.87 microg/ml. It also exhibited 65.5% and 98.2% of acetyl- and butyrylcholinesterase inhibition at 0.2 mg/ml, respectively. Compounds 2 and 8 showed the best potencies against P. falciparum W2 strain with IC(50) of 1.68 microM and 0.12 microM, (0.66 microg/ml and 0.054 microg/ml) respectively. All tested compounds showed good butyrylcholinesterase inhibition activities with compound 12 displaying the best potency (IC(50) 9.25+/-0.25 microM). All the tested compounds showed weak inhibitory activity against acetylcholinesterase.     

p190RhoGAP and Rap-dependent RhoGAP (ARAP3) inactivate RhoA in response to nerve growth factor leading to neurite outgrowth from PC12 cells

Rat pheochromocytoma (PC12) cells have been used to investigate neurite outgrowth. Nerve growth factor (NGF) has been well known to induce neurite outgrowth from PC12 cells. RhoA belongs to Ras-related small GTP-binding proteins, which regulate a variety of cellular processes, including cell morphology alteration, actin dynamics, and cell migration. NGF suppressed GTP-RhoA levels after 12 h in PC12 cells and was consistently required for a long time to induce neurite outgrowth. Constitutively active (CA)-RhoA suppressed neurite outgrowth from PC12 cells in response to NGF, whereas dominant-negative (DN)-RhoA stimulated it, suggesting that RhoA inactivation is essential for neurite outgrowth. Here, we investigated the mechanism of RhoA inactivation. DN-p190RhoGAP abrogated neurite outgrowth, whereas wild-type (WT)-p190RhoGAP and WT-Src synergistically stimulated it along with accelerating RhoA inactivation, suggesting that p190RhoGAP, which can be activated by Src, is a major component in inhibiting RhoA in response to NGF in PC12 cells. Contrary to RhoA, Rap1 was activated by NGF, and DN-Rap1 suppressed neurite outgrowth, suggesting that Rap1 is also essential for neurite outgrowth. RhoA was co-immunoprecipitated with Rap1, suggesting that Rap1 interacts with RhoA. Furthermore, a DN-Rap-dependent RhoGAP (ARAP3) prevented RhoA inactivation, abolishing neurite formation from PC12 cells in response to NGF. These results suggest that NGF activates Rap1, which, in turn, up-regulates ARAP3 leading to RhoA inactivation and neurite outgrowth from PC12 cells. Taken together, p190RhoGAP and ARAP3 seem to be two main factors inhibiting RhoA activity during neurite outgrowth in PC12 cells in response to NGF.     

The role of chalcones: helichrysetin,xanthohumol, and flavokawin-C in promoting neurite outgrowth in PC12 Adh cells

Chalcones are a group of compounds widely distributed in plant kingdom. The aim of this study was to assess the neurite outgrowth stimulatory activity of selected chalcones, namely helichrysetin, xanthohumol and flavokawin-C. Using adherent rat pheochromocytoma (PC12 Adh) cells, the chalcones were subjected to neurite outgrowth assay and the extracellular nerve growth factor (NGF) levels were determined. Xanthohumol (10 μg/mL) displayed the highest (p < 0.05) percentage of neurite-bearing PC12 Adh cells and the highest (p < 0.05) NGF level in the culture medium of xanthohumol-treated cells. While, helichrysetin induced a moderately high numbers of neurite-bearing cells, flavokawin-C did not stimulate neurite outgrowth. This work supports the potential use of xanthohumol as a potential neuroactive compound to stimulate neurite outgrowth.     

Sesquiterpenoids and flavonoids from the aerial parts of Tithonia diversifolia and their cytotoxic activity

Cytotoxicity-guided fractionation of the 80% EtOH extract of Tithonia diversifolia has resulted in the isolation of twelve sesquiterpenoids (1-12), including three new ones (4, 10, 12), and three known flavonoids (13-15). The structures of the new compounds were determined by analysis of their spectroscopic data. The isolated compounds showed cytotoxic activity against HL-60 leukemia cells with IC(50) values ranging from 0.13 to 13.0 microM, when etoposide used as a positive control gave an IC(50) value of 0.43 microM. The cancer growth inhibitory property of 9, the main cytotoxic compound in T. diversifolia, was examined using a disease-oriented panel composed of 39 human cancer cell lines in the Japanese Foundation for Cancer Research.