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A novel type of alpha,beta-butenolide alkaloid, uncinine (1), two novel oxoaporphines, artabonatine C (2) and artabonatine D (3), a new oxazoloaporphine, artabonatine E (4), and a new 7,7'-bisdehydroaporphine, artabonatine F (5), along with 25 known alkaloids, were isolated from Artabotrys uncinatus. The structures of 1-5 were determined using NMR and mass spectral data. Atherospermidine and squamolone exhibited cytotoxicity against hepatocarcinoma cancer cell lines (Hep G(2) and 2,2,15), and the activity of some of the alkaloids in an antithrombin assay is also discussed.

A novel alkaloid, hydrachine A (3), has been isolated, along with 15 known compounds, from the roots of Hydrangea chinensis. The structure and stereochemistry of the new alkaloid 3 was determined using extensive 2D NMR data.

Bioassay-directed fractionation of the stems of Rollinia mucosa led to the isolation of new N-methoxycarbonyl aporphine alkaloids, romucosine A (1), romucosine B (2), romucosine C (3), and romucosine D (4), along with the known alkaloid, N-methoxylcarbonyl-nornuciferine (5). Alkaloids 1 and 4 exhibited significant inhibition of collagen, arachidonic acid, and platelet activating factor-induced platelet aggregation, and alkaloid 3 also showed an inhibitory effect on arachidonic acid induced platelet aggregation.

Four alkaloids, annocherine A, annocherine B, cherianoine, and romucosine H, along with one known alkaloid, artabonatine B, were isolated from the MeOH extract of the stems of Annona cherimola. Their structures were identified on the basis of both analysis of their spectral data and from chemical evidence.

E2F-1 regulates the transcription of genes required for DNA synthesis. Previously, we have reported that UCN-01 suppresses E2F-1 protein expression without any noticeable effect on its mRNA level in gastric cancer cell line SK-GT5 (Clin. Cancer Res., 4: 2201-2206, 1998). In this study, we investigated the mechanism responsible for the suppression of E2F-1 expression by UCN-01 in SK-GT5 cells. After 24-h exposure to 1 microM UCN-01, E2F-1 protein expression was decreased by >99%. The suppressive effect of UCN-01 could be reversed by ubiquitin-dependent proteasome inhibitors such as calpain inhibitor I and lactacystin. Transfection experiments using expression plasmids encoding full-length E2F-1 or truncated E2F-1 with deletion of the COOH-terminal region (which is required for eliciting ubiquitination and protein degradation) revealed that the expression of truncated E2F-1 was not affected by UCN-01. Other cell-cycle-related and ubiquitin-proteasome-regulated proteins such as p21, p27, and cyclin B1 were not repressed by UCN-01 in E2F-1-overexpressing cells. In vitro-translated, full-length E2F-1 degraded more rapidly upon incubation with extracts from UCN-01-treated cells when compared with truncated E2F-1. Taken together, these data indicate that UCN-01 suppresses E2F-1 protein expression mediated by the ubiquitin-proteasome pathway in a specific manner.

A new halimane diterpene, 3beta,5beta, 16alpha-trihydroxyhalima-13(14)-en-15,16-olide (1), and a new oxoprotoberberine alkaloid, (-)-8-oxopolyalthiaine (2), along with 20 known compounds, were isolated from a methanolic extract of Polyalthia longifolia var. pendula. The structures of compounds 1 and 2 were established by spectroscopic analysis. Several of these compounds were evaluated for cytotoxicity toward a small panel of human cell lines.

Three new alkaloids, promucosine (1), romucosine F (2), and romucosine G (3), along with 28 known compounds, were isolated from the MeOH extract of stems of Annona purpurea. The structures of 1-3 were determined on the basis of spectral data and chemical evidence.

Bioactivity-directed fractionation led to the isolation of two new 7-dehydroaporphine alkaloids, 7-hydroxy-dehydrothalicsimidine (1) and 7-formyl-dehydrothalicsimidine (2), along with the five known alkaloids, thalicsimidine (3), norpurpureine (4), N-methyllaurotetanine (5), lirinidine (6) and N-methylasimilobine (7), from the leaves of Annona purpurea. Structural elucidation of these compounds was established by mass and spectroscopic analyses. Among them, 1, 3, 4, 6 and 7 exhibited significant inhibition of collagen, arachidonic acid and platelet activating factor-induced platelet aggregation; 1 also showed inhibition against thrombin-induced platelet aggregation.

Bioactivity-directed fractionation led to the isolation of 19 compounds, including three oxoaporphines, oxopurpureine (5), oxonuciferine (6), and oxoglaucine (7); three aporphines, (+)-predicentrine (8), (-)-glaucine (9), and thalbaicalidine (10); one aporphine sensu stricto, N-formyl-purpureine (11); one proaporphine, glaziovine; one phenanthrene, thalicpureine (12); two 6a,7-dehydroaporphines, dehydrolirinidine (13) and 7-hydroxy-dehydroglaucine (14); four flavonoids, quercetin-3-O-rhamnoside, kaempferol-3-O-rhamnoside, isorhamnetin-3-O-rhamnoside, and tanarixetin-3-O-rhamnoside; one purine, adenine; one lactam amide, squamolone; and two steroids, beta-sitosterol and beta-sitosterol-beta-D-glucoside from the MeOH extract of the leaves of Formosan Annona purpurea. Among them, 11-14 were characterized as new compounds and alkaloids, 5-8, 10, and 12-14 exhibited significant antiplatelet aggregation activity.

Using a bioassay-directed fractionation method, three new compounds, including an aporphine alkaloid, cassyformine (4); an oxoaporphine alkaloid, filiformine (8), and a lignan, (+)-diasyringaresinol (10), along with 14 known compounds, were further isolated and characterized from the MeOH extract of the fresh herbs of Cassytha filiformis. Among the isolates of this plant, cathafiline (1), cathaformine (2), actinodaphnine (3), N-methylactinodaphnine (5), predicentrine (6), and ocoteine (7) exhibited significant antiplatelet aggregation activity.

Using antiplatelet aggregation as a guide to fractionation, seven aporphines, actinodaphnine (1), N-methylactinodaphnine (2), launobine (3), dicentrine (4), O-methylbulbocapnine (5), hernovine (7), and bulbocapnine (9), and two oxoaporphines, dicentrinone (6) and liriodenine (8), were isolated from the stems of Illigera luzonensis. Among them, compounds 2, 4, 5, 8, and 9 were isolated for the first time from this species. Moreover, compounds 1-5, and 8 showed significant antiplatelet aggregation and compounds 1 and 6 exhibited significant vasorelaxant activities, respectively.

Members of a series of benzylisoquinoline and phenanthrene alkaloids were tested for their antiplatelet and vasorelaxing actions. Atherosperminine HClO4 (15) and atherosperminium I (16) showed strong inhibition of adenosine 5'-diphosphate-induced platelet aggregation. dl-N-Methylcoclaurine (1), dl-coclaurine (6), 15, 16, xylopine hydroxylamine (18), and atherosperminine N-oxide (19) showed strong inhibition of arachidonic acid-induced platelet aggregation. Compounds 1, 15, 16, dicentrine methine (17), 18, and 19 showed strong inhibition of collagen-induced platelet aggregation. d-(-)-Magnocurarine I (8), 15, and 16 showed strong inhibition of platelet-activating factor-induced platelet aggregation. Compounds 15, 17, and 19 showed vasorelaxing action in rat thoracic aorta.

The pharmacological activity of atherosperminine, isolated from Fissistigma glaucescens, was determined in isolated guinea-pig trachealis. Atherosperminine (25-100 microM) and theophylline (10-1000 microM) both inhibited the contractile response caused by carbachol, prostaglandin F2 alpha (PGF2 alpha), U46619 (thromboxane A2 analogue), leukotriene C4 (LTC4) and Ca2+ (in the presence of 120 mM KCl) in a concentration-dependent manner. The inhibition was characterized by a rightwards shift of the concentration-response curves with suppression of the maximal contraction. Propranolol (1 microM), glibenclamide (10 microM) and removal of tracheal epithelium did not modify the relaxant action of atherosperminine. Atherosperminine (25 and 50 microM) caused a 2.4- and 5.0-fold, respectively, potentiation of the action of forskolin to cause tracheal relaxation but did not potentiate the action of sodium nitroprusside or cromakalim. Atherosperminine (50 microM) potentiated the action of forskolin to increase tissue cAMP content and, in higher concentrations (100 and 250 microM), itself increased tissue cAMP but not cGMP content. Atherosperminine markedly inhibited cAMP phosphodiesterase but not cGMP phosphodiesterase in homogenates of guinea-pig trachealis. It is concluded that atherosperminine exerts a non-specific relaxant effect on the trachealis. Its major mechanism of action appears to be inhibition of cAMP phosphodiesterase, perhaps with a minor effect on cGMP phosphodiesterase at higher concentrations.

The pharmacological properties and mechanism of the action of protopine on isolated rat thoracic aorta were examined. It inhibited norepinephrine (NE, 3 microM)-induced tonic contraction in rat thoracic aorta in a concentration-dependent manner (25-100 micrograms/ml). The phasic contraction caused by NE was inhibited only by a high concentration of protopine (100 micrograms/ml). At the plateau of NE-induced tonic contraction, the addition of protopine also caused relaxation. This relaxing effect of protopine was not antagonized by indomethacin (20 microM) or methylene blue (50 microM), and it still existed in denuded rat aorta or in the presence of nifedipine (2-100 microM). Protopine also inhibited high potassium (60 mM)-induced, calcium-dependent (0.03-3 mM) contraction of rat aorta in a concentration-dependent manner. Neither cAMP nor cGMP level was changed by protopine. Both the formation of inositol monophosphate caused by NE and the phasic contraction induced by caffeine were also not affected by protopine. 45Ca2+ influx caused by either NE or K+ was inhibited by protopine concentration-dependently. It is concluded that protopine relaxed the rat thoracic aorta mainly by suppressing the Ca2+ influx through both voltage- and receptor-operated calcium channels.

Bioactivity-guided fractionation of a CHCl3 extract of the leaves of Piper aborescens afforded a new cytotoxic pyridone alkaloid, N-(3-methoxy-4,5-methylenedioxydihydrocinnamoyl)-delta 3-pyridin-2-one [1], as well as three known cytotoxic pyridone alkaloids, N-(3-methoxy-4,5-methylenedioxycinnamoyl)-delta 3-pyridin-2-one [2], piplartine [3], and piplartine dimer A [4].

The stem and stem bark of Polyalthia longifolia afforded the cytotoxic aporphine alkaloid liriodenine [1], as well as two aporphine alkaloids, noroliveroline [2] and oliveroline-beta-N-oxide [3] and three azafluorene alkaloids, darienine [4], polyfothine [6], and isooncodine [7], which are not bioactive. Polyfothine [6] and isooncodine [7] are new natural compounds.

A number of benzylisoquinoline alkaloids of plants indigenous to Taiwan were isolated and purified and their chemical structures were determined. These compounds, namely benzyltetrahydroisoquinolines or their N-oxides, aporphines, oxoaporphines and 1-N, N-dimethylaminoethylphenanthrenes or their N-oxides, were studied for their potency in inhibiting precursor incorporation into DNA, RNA and protein. Inhibition of murine leukemic L1210 and human leukemic CCRF-CEM and HL-60 cell growth was also examined. Of the compounds evaluated for cell growth inhibition, (+-)-N-methylcoclaurine(3), (-)-norannuradhapurine HBr(19), oxoglaucine (23), dicentrine methine (27) and 1-(N-methyl-N-hydroxyaminoethyl)-3, 4-methylenedioxy-7-methoxyphenanthrene (28) showed an IC50 less than 10 microM.

A series of 53 isoquinoline alkaloids and their N-oxides have been tested for their cytotoxicity against A-549, HCT-8, KB, P-388, and L-1210 cells. These alkaloids include two tetrahydroprotoberberines, two protoberberines, six aporphines, one morphinandienone, five oxoaporphines, seven phenanthrenes, one spirobenzylisoquinoline N-oxide, nine aporphine N-oxides, seven benzyltetrahydroisoquinoline N-oxides, one benzylisoquinoline N-oxide, one protopine N-oxide, three tetrahydroprotoberberine N-oxides, four pavine N-oxides, and four phenanthrene N-oxides. The results are discussed on the basis of structure-activity relationships.

Protopine inhibited the aggregation and ATP release of rabbit platelets induced by ADP, arachidonic acid, PAF, collagen and ionophore A23187. Although the platelet aggregation caused by thrombin was not inhibited by protopine (100 micrograms/ml), the release reaction was partially suppressed. In rabbit platelet-rich plasma, protopine also inhibited the platelet aggregation caused by ADP, arachidonic acid, PAF and collagen. The thromboxane B2 formation of washed platelets caused by arachidonic acid, collagen, ionophore A23187 and thrombin was suppressed by protopine. Protopine inhibited the intracellular calcium increase caused by arachidonic acid in quin-2/AM loaded rabbit platelets. In the presence of indomethacin, the intracellular calcium increase caused by collagen and PAF was completely suppressed by protopine, and the intracellular calcium increase caused by thrombin was partially inhibited. The phosphoinositides breakdown caused by collagen and PAF was inhibited by protopine, but that by thrombin was not affected significantly. Protopine did not cause the elevation of cyclic AMP level of platelets. It is concluded that the antiplatelet effects of protopine is due to inhibition on thromboxane formation and phosphoinositides breakdown and then lead to the decrease of intracellular calcium concentration.