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A new ent-kaurene diterpene, pseudoirroratin A (1), and a known diterpene, pseurata A, were isolated from Isodon pseudo-irrorata. Compound 1 showed significant cytotoxicity against the Lu1, SW626, LNCaP, KB, and HOS cancer cell lines with IC(50) values of 0.26 (0.75), 0.20 (0.57), 0.90 (2.59), 0.90 (2.59), and 0.50 (1.44) microg/mL (microM), respectively. The structure of 1 was elucidated by spectroscopic means including 1D and 2D NMR techniques.

The eudesmane sesquiterpenoid, verticillatol (1), as well as the lignan, (+)-5'-demethoxyepiexcelsin (2), and a known lignan, (+)-epiexcelsin (3), were isolated from Litsea verticillata Hance. Lignan 2 showed moderate anti-HIV activity with an IC(50) value of 16.4 microg/ml (42.7 microM), while the known lignan 3 was inactive up to a concentration of 20 microg/ml (48.3 microM). Compound 1 demonstrated weak activity with an IC(50) value of 34.5 microg/ml (144.7 microM) while being devoid of cytotoxicity at 20 microg/ml. The structures were elucidated by 1D and 2D NMR spectroscopy, and the absolute configuration of the new sesquiterpenoid was determined by the generation of Mosher esters.

Fractionation of an ethyl acetate-soluble extract of the bark of Artocarpus dadah has led to the isolation of three new prenylated stilbenoid derivatives, 3-(gamma,gamma-dimethylallyl)resveratrol (1), 5-(gamma,gamma-dimethylallyl)oxyresveratrol (2), 3-(2,3-dihydroxy-3-methylbutyl)resveratrol (3), and a new benzofuran derivative, 3-(gamma,gamma-dimethylpropenyl)moracin M (4), along with six known compounds, oxyresveratrol, (+)-catechin, afzelechin-3-O-alpha-L-rhamnopyranoside, (-)-epiafzelechin, dihydromorin, and epiafzelechin-(4beta-->8)-epicatechin. From an ethyl acetate-soluble extract of the twigs of the same plant were isolated compound 4 and two new neolignan derivatives, dadahols A (5) and B (6), as well as 10 known compounds, oxyresveratrol, (+)-catechin, afzelechin-3-O-alpha-L-rhamnopyranoside, resveratrol, steppogenin, moracin M, isogemichalcone B, gemichalcone B, norartocarpetin, and engeletin. The structures of compounds 1-6 were determined using spectroscopic and chemical methods. Isolates were evaluated for their inhibitory effects against both cyclooxygenase-1 (COX-1) and -2 (COX-2) and in a mouse mammary organ culture assay.

Bioassay-guided fractionation of an ethyl acetate-soluble extract from the whole plants of Broussonetia papyrifera, using an in vitro aromatase inhibition assay, led to the isolation of five new active compounds, 5,7,2',4'-tetrahydroxy-3-geranylflavone (1), isogemichalcone C (8), 3'-[gamma-hydroxymethyl-(E)-gamma-methylallyl]-2,4,2',4'-tetrahydroxychalcone 11'-O-coumarate (9), demethylmoracin I (10), and (2S)-2',4'-dihydroxy-2' '-(1-hydroxy-1-methylethyl)dihydrofuro[2,3-h]flavanone (11), and 10 known (12-21) compounds which were also found to be active. Of these compounds, the most potent were 9 (IC(50) 0.5 microM), 11 (IC(50) 0.1 microM), isolicoflavonol (12, IC(50) 0.1 microM), and (2S)-abyssinone II (13, IC(50) 0.4 microM). Additionally, six new compounds, 5,7,3',4'-tetrahydroxy-6-geranylflavonol (2), 5,7,3',4'-tetrahydroxy-3-methoxy-6-geranylflavone (3), (2S)-7,4'-dihydroxy-3'-prenylflavan (4), 1-(2,4-dihydroxyphenyl)-3-(4-hydroxyphenyl)propane (5), 1-(2,4-dihydroxy-3-prenylphenyl)-3-(4-hydroxyphenyl)propane (6), and 1-(4-hydroxy-2-methoxyphenyl)-3-(4-hydroxy-3-prenylphenyl)propane (7), were isolated and characterized, but proved to be inactive as aromatase inhibitors, as were an additional 21 known compounds. The structures of the new compounds (1-11) were elucidated by spectroscopic methods. Structure-activity relationships in the aromatase assay were determined for the benzofurans, biphenylpropanoids, coumarins, and various types of flavonoids (chalcones, flavans, flavanones, and flavones) obtained among a total of 42 constituents of B. papyrifera.

Further investigation on the alcohol extract of the barks of Taxus yunnanensis led to the isolation of four taxoids, namely, 7beta-xylosyl-taxol D, taxuyunnanines P, Q and R, along with the known taxuyuntin G (2). Four are rearranged taxoids with an 11(15-->1)-abeotaxoid skeleton and an opened oxetane ring moiety. Structures were determined by spectroscopic and chemical means.

[structure: see text] Aiphanol (1), a novel stilbenolignan, along with isorhapontigenin (2), piceatannol (3), and luteolin, were isolated by bioassay-guided fractionation from the seeds of Aiphanes aculeata Willd. (Arecaceae). The structure of compound 1 was elucidated by spectroscopic methods. Compound 1 is based on an unprecedented stilbenolignan skeleton in which a stilbene moiety is linked with a phenylpropane unit through a dioxane bridge. Compounds 1 and 2 exhibited significant inhibitory activities against cyclooxygenases-1 and -2.

Two new compounds, (6S)-hydroxy-29-nor-3,4-seco-cycloart-4(30),24-dien-3-oic acid (1) and 8-[1-(3,4-dihydroxyphenyl)-3-methoxy-3-oxopropyl]epicatechin (3), were isolated by bioassay-guided fractionation from the aerial parts of Antirhea acutata (DC.) Urb. (Rubiaceae). Compound 1 showed moderate inhibitory activities in cyclooxygenase-1 and -2 assays (IC(50) 43.7 and 4.7 microM, respectively), while compound 3 was active in 1,1-diphenyl-2-picrylhydrazyl free-radical and cytochrome c reduction antioxidant assays (IC(50) 29.1 and 16.3 microM, respectively). Additionally, one further new compound was isolated, (3S,24S)-25-trihydroxy-9,19-cycloartane-29-oic acid (2), but this was inactive in the bioassay systems used. Compound 1 is based on the unprecedented 29-nor-3,4-seco-cycloartane skeleton.

Bioassay-directed fractionation led to the isolation of 14 compounds, six of which possess antimalarial activity, from the dried leaves and stems of Rhaphidophora decursiva. Polysyphorin (1) and rhaphidecurperoxin (6) showed strong activities against Plasmodium falciparum. Rhaphidecursinol A (2), rhaphidecursinol B (3), grandisin (4), and epigrandisin (5) were less active against the same organism. Among the isolates, rhaphidecursinol A (2) and rhaphidecursinol B (3) were determined to be new neolignans, and rhaphidecurperoxin (6) is a new benzoperoxide. Known compounds isolated include polysyphorin (1), grandisin (4), epigrandisin (5), (+)-medioresinol, (-)-pinoresinol, (-)-syringaresinol, (+)-glaberide I, (+)-dehydrovomifoliol, (-)-liliolide, (-)-hydroxydihydrobovolide, and N-butylbenzamide, of which compound 1 appears worthy of further evaluation as an antimalarial agent. Structure elucidation and identification were accomplished by spectroscopic means including 1D and 2D NMR analyses.

Eight botanical preparations that are commonly used for the treatment of menopausal symptoms were tested for estrogenic activity. Methanol extracts of red clover (Trifolium pratense L.), chasteberry (Vitex agnus-castus L.), and hops (Humulus lupulus L.) showed significant competitive binding to estrogen receptors alpha (ER alpha) and beta (ER beta). With cultured Ishikawa (endometrial) cells, red clover and hops exhibited estrogenic activity as indicated by induction of alkaline phosphatase (AP) activity and up-regulation of progesterone receptor (PR) mRNA. Chasteberry also stimulated PR expression, but no induction of AP activity was observed. In S30 breast cancer cells, pS2 (presenelin-2), another estrogen-inducible gene, was up-regulated in the presence of red clover, hops, and chasteberry. Interestingly, extracts of Asian ginseng (Panax ginseng C.A. Meyer) and North American ginseng (Panax quinquefolius L.) induced pS2 mRNA expression in S30 cells, but no significant ER binding affinity, AP induction, or PR expression was noted in Ishikawa cells. Dong quai [Angelica sinensis (Oliv.) Diels] and licorice (Glycyrrhiza glabra L.) showed only weak ER binding and PR and pS2 mRNA induction. Black cohosh [Cimicifuga racemosa (L.) Nutt.] showed no activity in any of the above in vitro assays. Bioassay-guided isolation utilizing ER competitive binding as a monitor and screening using ultrafiltration LC-MS revealed that genistein was the most active component of red clover. Consistent with this observation, genistein was found to be the most effective of four red clover isoflavones tested in the above in vitro assays. Therefore, estrogenic components of plant extracts can be identified using assays for estrogenic activity along with screening and identification of the active components using ultrafiltration LC-MS. These data suggest a potential use for some dietary supplements, ingested by human beings, in the treatment of menopausal symptoms.

The pregnane steroid, (E)-aglawone, along with four known triterpenes, and a known sterol mixture were isolated from the bark of Aglaia lawii (Wight) Saldanha ex Ramamoorty (Meliaceae). The structural determination/identification was accomplished by a combination of 1D- and 2D-NMR spectroscopic techniques. The relative stereochemistry of the known triterpene, 20S,24S-epoxydammarane-3alpha,25-diol acetate, was also unequivocally determined for the first time by X-ray crystallography. The isolates were not active against various human cancer cell lines.

Six constituents (1-6) were isolated from EtOAc-soluble partitions of two separate collections of the whole plants of Cotinus coggygria, namely, disulfuretin ¿2,2'-[1,2-bis(3,4-dihydroxyphenyl)-1, 2-ethanedylidene]bis[6-hydroxy-3(2H)-benzofuranone] (1)¿, sulfuretin (2), sulfurein (3), gallic acid (4), methyl gallate (5), and pentagalloyl glucose (6). The structure of the novel biaurone 1 was determined by spectral and chemical methods. Compounds 1-6 were found to be potent antioxidants in a 1,1-diphenyl-2-picrylhydrazyl free-radical scavenging assay.

Five new 11(15-->1)-abeo-taxane diterpenoids, taxuyunnanines K-O (1-5), were isolated from an ethanol extract of the bark of Taxus yunnanensis, and their structures were determined using MS and NMR techniques. Compounds 1/2 and 4/5 are rearranged taxane diterpenoids possessing an opened oxetane ring moiety at C4(20). Compounds 4/5 are rearranged taxoids lacking an oxygenated functionality at C-4.

A liquid chromatography-tandem mass spectrometry (LC-MS-MS) method was developed to distinguish Asian ginseng (Panax ginseng C. A. Meyer) and North American ginseng (Panax quinquefolius L.). The method is based on the baseline chromatographic separation of ginsenoside Rf and 24(R)-pseudoginsenoside F11, two potential chemical markers present in ginseng root methanolic extracts, and their unambiguous on-line identification using tandem mass spectrometry. Consistent with the literature, 24(R)-pseudoginsenoside F11 was detected in abundance in North American ginseng roots in excess of 0.1% (w/w) of the dried root. In contrast to some reports, 24(R)-pseudoginsenoside F11 was also identified in Asian ginseng roots at trace levels using LC-MS-MS but at less than 0.0001% (w/w). Besides showing identical tandem mass spectra to authentic 24(R)-pseudoginsenoside F11, the corresponding compound in Asian ginseng root coeluted with standard under different HPLC conditions, thus confirming this compound as 24(R)-pseudoginsenoside F11. Another ginsenoside often used to distinguish Asian and North American ginseng, ginsenoside Rf, was found in abundance in Asian ginseng roots at more than 0.021% (w/w). In Asian ginseng roots, the ratio of ginsenoside Rf to 24(R)-pseudoginsenoside F11 exceeded 700:1. The limit of detection of ginsenoside Rf or 24(R)-pseudoginsenoside F11 was 120 pg injected on-column, and the limit of quantification was 240 pg on-column. In summary, LC-MS-MS analysis of ginseng products for the presence and ratio of ginsenoside Rf and 24(R)-pseudoginsenoside F11 may be used for the unambiguous identification of Asian and North American ginsengs.

Two dimeric naphthoquinones, diospyrin and isodiospyrin, isolated from the root of Diospyros piscatoria (Gurke), a common ingredient in several folk medicines, have been shown to have a broad spectrum of antibacterial activity. The minimum inhibitory concentrations (MICs) of diospyrin against Streptococcus pyogenes ATCC 12344 and Streptococcus pneumoniae ATCC 33400 ranged from 1.56 to 50 microg/mL. While those against Salmonella choleraesuis serotype typhi (S. typhi), ATCC 6539 and Mycobacterium chelonae ATCC 19977 were between 25 and 100 microg/mL. Isodiospyrin was more active than its racemic isomer diospyrin. The MICs against Gram-positive bacteria ranged from 0.78 to 50 microg/mL. While those against Pseudomonas aeruginosa ATCC 15443 and S. typhi ranged from 50 to 100 microg/mL. The MIC for M. chelonae was between 6.25 and 25 microg/mL. MICs were found to increase with the concentration of cells used for the inoculum. The MICs for Bacillus subtilis ATCC 6633 increased up to the highest concentration of cells tested. The same phenomenon was observed on M. chelonae, but with better effect in the latter. The kinetics of bacteria studies against both B. subtilis and M. chelonae increases with increasing concentration of isodiospyrin tested. Two tetrameric forms of plumbagin were isolated. The naphthoquinone bisisodiospyrin, gave MIC values between 300 and 400 micro g/mL. The second, as yet unidentified tetramer, was not active at 500 micro g/mL., (Copyright 2000 John Wiley & Sons, Ltd.)

From the leaves of Isodon megathyrsus, a novel ent-kaurene diterpene, megathyrin B, was isolated and its structure determined as 1 alpha,7 beta,11 beta,15 beta-tetrahydroxy-ent-7 alpha,20-epoxy-kaur-16-ene by 1D- and 2D-NMR spectral analysis. Additionally, its stereochemistry was unambiguously assigned by X-ray crystallography. This compound was cytotoxic to the KB and KB-V cell lines.

Four novel steroidal alkaloids, (+)-(20S)-20-(dimethylamino)-3-(3'alpha-isopropyl)-lactam-5alpha-+ ++preg n-2-en-4-one (1), (+)-(20S)-20-(dimethylamino)-16alpha-hydroxy-3-(3'alpha-isopropyl) -la ctam-5alpha-pregn-2-en-4-one (2), (+)-(20S)-3-(benzoylamino)-20-(dimethylamino)-5alpha-pregn-2-en-++ +4beta -yl acetate (3), and (+)-(20S)-2alpha-hydroxy-20-(dimethylamino)-3beta-phthalimido-5 alpha- pregnan-4beta-yl acetate (4), as well as five known compounds, (-)-pachyaximine A (5), (+)-spiropachysine (6), (+)-axillaridine A (7), (+)-epipachysamine D (8), and (+)-pachysamine B (9), were isolated from Pachysandra procumbens, using a bioassay-guided fractionation based on inhibition of 3H-tamoxifen binding at the antiestrogen binding site (AEBS). Compounds 1-7 and 9 demonstrated significant activity as AEBS-inhibitory agents, and compounds 3, 5 and 9 were found to potentiate significantly the antiestrogenic effect mediated by tamoxifen in cultured Ishikawa cells. The structure elucidation of compounds 1-4 was carried out by spectral data interpretation.

Starting with an extract derived from the stem of Macleaya cordata (Papaveraceae) that was active in the process of inhibiting phorbol 12,13-dibutyrate binding to partially purified protein kinase C (PKC), the benzophenanthridine alkaloid angoline was isolated and identified. This discovery appeared in context, as a related benzophenanthridine alkaloid, chelerythrine, has been reported to mediate a variety of biological activities, including potent and selective inhibition of protein kinase C (PKC). However, in our studies, angoline was not observed to function as a potent inhibitor of PKC. Moreover, we were unable to confirm the reported inhibitory activity of chelerythrine. In a comprehensive series of studies performed with various PKC isozymes derived from a variety of mammalian species, neither chelerythrine nor angoline inhibited activity with high potency. To the contrary, chelerythrine stimulated PKC activity in the cytosolic fractions of rat and mouse brain in concentrations up to 100 microM. In addition, chelerythrine and angoline did not inhibit [3H]phorbol 12,13-dibutyrate binding to the regulatory domain of PKC at concentrations up to 40 microg/ml, and no significant alteration of PKC-alpha, -beta, or -gamma translocation was observed with human leukemia (HL-60) cells in culture. Further, chelerythrine did not inhibit 12-O-tetradecanoylphorbol 13-acetate-induced ornithine decarboxylase activity with cultured mouse 308 cells, but angoline was active in this capacity with an IC50 value of 1.0 microg/ml. A relatively large number of biological responses have been reported in studies conducted with chelerythrine, and alteration of PKC activity has been considered as a potential mechanism of action. In light of the current report, mechanisms independent of PKC inhibition should be considered as responsible for these effects.

A novel flavonoid diglycoside, 5,6,7,4'-tetrahydroxyflavonol 3-O-rutinoside (1), and a previously known compound, kaempferol 3-O-neohesperidoside (2), were isolated from an ethyl acetate extract of Daphniphyllum calycinum leaves that showed significant activity in a 1,1-diphenyl-2-picrylhydrazyl (DPPH) free-radical assay. The structure of 1 was elucidated by a combination of spectroscopic methods, and compounds 1 and 2 were found to be moderately active as antioxidants in the DPPH assay.

Utilizing the hepatitis B virus (HBV)-producing hepatoblastoma cell line, HepG2.2.15, which is stably transfected with the cloned HBV genome, methods were devised to examine the effects of test substances on intracellular extrachromosomal HBV DNA levels and secretion of hepatitis B surface antigen (HBsAg). The known inhibitor of HBV replication, dideoxycytosine (ddC), had a minor effect on the secretion of HBsAg, but >90% of intracellular extrachromosomal HBV DNA expression was lost at a non-cytotoxic drug concentration (25μM). This inhibitory effect was reversed when ddC was removed from the medium. Of 19 plant materials tested, extracts from the aerial parts of Clematis sinensis Lour, and Clerodendron inerme R. Br. significantly inhibited the secretion of HBsAg into the culture medium at non-cytotoxic concentrations, but had no effect on intracellular extrachromosomal HBV DNA levels. This system is useful for the evaluation of test materials, or combinations of test materials, for their potential to inhibit HBV markers., (Copyright © 1996 Gustav Fischer Verlag. Published by Elsevier GmbH.. All rights reserved.)

Resveratrol, a phytoalexin found in grapes and other food products, was purified and shown to have cancer chemopreventive activity in assays representing three major stages of carcinogenesis. Resveratrol was found to act as an antioxidant and antimutagen and to induce phase II drug-metabolizing enzymes (anti-initiation activity); it mediated anti-inflammatory effects and inhibited cyclooxygenase and hydroperoxidase functions (antipromotion activity); and it induced human promyelocytic leukemia cell differentiation (antiprogression activity). In addition, it inhibited the development of preneoplastic lesions in carcinogen-treated mouse mammary glands in culture and inhibited tumorigenesis in a mouse skin cancer model. These data suggest that resveratrol, a common constituent of the human diet, merits investigation as a potential cancer chemopreventive agent in humans.

A novel lignan, guaiacylglycerol-beta-O-6'-(2-methoxy)cinnamyl alcohol either, three known simaroubolides, brusatol, dehydrobrusatol, yadanziolide C, and the known terpenoid, blumenol A, were obtained as active compounds from an ethyl acetate-soluble extract of Brucea javanica, using a bioassay based on the induction of cell differentiation with human promyelocytic leukemia (HL-60) cells. Also obtained were the known coumarinolignan, cleomiscosin A, and the known quassinoid glycoside, bruceoside B, which were inactive in the HL-60 cell test system. The structure of the new lignan was determined by a combination of 1D and 2D NMR techniques.

In a continuation of phytochemical studies on the underground organs of Liriope spicata var. prolifera, four new steroidal glycosides, lirioproliosides A-D, along with two known compounds, 25(S)-ruscogenin 1-O-[alpha-L-rhamnopyranosyl(1-->2)[beta-D-xylopyranosyl (1-->3)]-beta-D-fucopyranoside and ophiopogonin A, were identified. The structures of lirioproliosides A-D were established by a combination of spectroscopic and chemical methods as 25(S)-ruscogenin 1-O-[alpha-L-rhamnopyranosyl(1-->2)][beta-D-xylopyranosyl (1-->3)]-beta-D-fucopyranoside-3-O-alpha-L-rhamnopyranoside, 25(S)-ruscogenin 1-O-[3-O-acetyl-alpha-L-rhamnopyranosyl(1-->2)]-beta-D-fucopyranoside, 25(S)-ruscogenin (1-O-[2-O-acetyl-alpha-L-rhamnopyranosyl (1-->2)]-beta-D-fucopyranoside and ruscogenin (1-O-[2-O-acetyl-alpha-L-rhamnopyranosyl(1-->2)]-beta-D-fucopyranoside, respectively. Among these steroidal glycosides, ophiopogonin A and lirioprolioside B, and lirioproliosides C and D, were isolated as epimeric pairs.

The mammalian lignans enterodiol (ED) and enterolactone (EL) produced from colonic bacterial action on dietary precursors have exhibited anticarcinogenic effects in vitro. The major lignan precursor in flaxseed (a rich source) has been identified as secoisolariciresinol diglycoside (SDG). The purpose of this study was to first isolate SDG and determine whether 1) SDG accounted for all the lignan production from flaxseed; 2) this production was dose-related; and 3) a relationship between in vitro production and in vivo urinary excretion existed. Extraction of flaxseed with dioxane:ethanol (1:1, v/v) followed by chromatographic separations yielded the purified SDG. Rats were fed a high fat diet without/with 2.5, 5 or 10 g/100g ground flaxseed or 1.1, 2.2 or 4.4 micromol SDG/d (equivalent to levels in the respective flaxseed diets) for 4 wk. In vitro lignan production was assessed by fermenting flaxseed or SDG for 24 h with human fecal inoculum. Urinary lignan excretion increased linearly with doses from 0-5% flaxseed and 0-2.2 micromol SDG/d followed by a plateau, indicating a threshold response. When all doses were considered, a curvilinear relationship was observed. A similar trend was seen in vitro for SDG, resulting in a high correlation between in vitro production and in vivo excretion of lignans (r = 0.990, P < 0.02). Thus in vivo response can be predicted with confidence based on in vitro results. Theoretical urinary ED + EL from the SDG present in flaxseed correlated with the actual excretion in flaxseed-fed animals (r = 0.655, P < 0.005). However, urinary ED + EL of SDG-fed rats was only 20% of levels of flaxseed-fed rats, indicating the presence of other precursors or incomplete conversion of SDG to ED and EL.

An A ring-secocycloartene triterpenoid, nigranoic acid (3,4-secocycloarta-4(28),24-(Z)-diene-3,-26-dioic acid, (1) was isolated from the stems of Schisandra sphaerandra, a Chinese traditional medicinal plant. Its structure elucidation and unambiguous NMR spectral assignment were achieved by the combination of 1D- and 2D-NMR techniques with the aid of computer modeling. Nigranoic acid showed activity in several anti-HIV reverse transcriptase and polymerase assays.

Secoisolariciresinol diglycoside (SD), a mammalian lignan precursor found in high-fiber foods, was isolated from flaxseed and tested for effects on mammary tumorigenesis in rats fed a high-fat (20%) diet. Ingestion of purified SD at 1.5 mg/day for 20 weeks starting 1 week after treatment with the carcinogen dimethylbenzanthracene resulted in a 37% reduction (p < 0.05) in the number of tumors per tumor-bearing rat and a 46% reduction (p < 0.05) in the number of tumors per tumor-bearing rat and a 46% reduction (p < 0.05) in the number of tumors per number of rats in each group. Urinary mammalian lignan excretion significantly increased (p < 0.0001) with SD treatment, indicating the conversion of SD to mammalian lignans. No enlargement or gross abnormalities of the major organs were observed in the SD-treated rats. This study showed, for the first time, that SD has an antitumor effect when provided at the early promotion stage of tumorigenesis and may contribute to the health benefits of high-fiber foods.

As a result of bioassay-guided fractionation, betulinic acid, a pentacyclic triterpene, was identified as a melanoma-specific cytotoxic agent. In follow-up studies conducted with athymic mice carrying human melanomas, tumour growth was completely inhibited without toxicity. As judged by a variety of cellular responses, antitumour activity was mediated by the induction of apoptosis. Betulinic acid is inexpensive and available in abundant supply from common natural sources, notably the bark of white birch trees. The compound is currently undergoing preclinical development for the treatment or prevention of malignant melanoma.

Swertifrancheside [1], a new flavonone-xanthone glucoside isolated from Swertia franchetiana, 1 beta-hydroxyaleuritolic acid 3-p-hydroxybenzoate [2], a triterpene isolated from the roots of Maprounea africana, and protolichesterinic acid [3], an aliphatic alpha-methylene-gamma-lactone isolated from the lichen Cetraria islandica, were found to be potent inhibitors of the DNA polymerase activity of human immunodeficiency virus-1 reverse transcriptase (HIV-1 RT), with 50% inhibitory doses (IC50 values) of 43, 3.7, and 24 microM, respectively. They were not cytotoxic with cultured mammalian cells. The kinetic mechanisms by which compounds 1-3 inhibited HIV-1 RT were studied as was their potential to inhibit other nucleic acid polymerases. Swertifrancheside [1] bound to DNA and was shown to be a competitive inhibitor with respect to template-primer, but a mixed-type competitive inhibitor with respect to TTP. On the other hand, 1 beta-hydroxyaleuritolic acid 3-p-hydroxybenzoate [2] and protolichesterinic acid [3] were mixed-type competitive inhibitors with respect to template-primer and noncompetitive inhibitors with respect to TTP. Therefore, the mechanism of action of 1 beta-hydroxyaleuritolic acid 3-p-hydroxybenzoate [2] and protolichesterinic acid [3] as HIV-1 RT inhibitors involves nonspecific binding to the enzyme at nonsubstrate binding sites, whereas swertifrancheside [1] inhibits enzyme activity by binding to the template-primer.

Terminal differentiation of human promyelocytic leukemia (HL-60) cells can be induced by a variety of chemical agents and this process can be monitored readily by the generation of morphologically, histochemically, and functionally mature granulocytes and monocytes/macrophages. The availability of this model has heightened interest in the possible therapeutic role of inducers of myeloid differentiation for the treatment of leukemia and other neoplasms. In addition, however, potent cancer chemopreventive agents induce HL-60 cell differentiation at very low dose levels. Thus, as part of our search for natural product chemopreventive agents, extracts derived from nearly 400 plants were tested for their potential to induce HL-60 cell differentiation. As a result, 17 plant extracts were judged to be active (ED50 values < or = 4 micrograms/ml). One of most potent leads was an extract derived from Dirca occidentalis Gray (Thymelaeaceae) (ED50, 0.14 micrograms/ml), and bioassay-guided fractionation led to the identification of genkwanin (I), (+/-)-lariciresinol (II) and sitoindoside II (IV) as active principles, with ED50 values of 18.3, 1.1 and 0.069 microM, respectively. Based on these data, we conclude that the HL-60 cell differentiation system is a valid and useful model for the discovery of natural product cancer chemopreventive or chemotherapeutic agents.

For the discovery of new cancer chemopreventive agents, we have studied the potential of plant extracts to inhibit phorbol ester-induced ornithine decarboxylase (ODC) activity in cell culture. Four active rotenoids were obtained from the African plant Mundulea sericea (Leguminosae). These isolates were highly potent when evaluated for inhibition of chemically induced preneoplastic lesions in mammary organ culture and inhibition of papillomas in the two-stage mouse skin model, and they appear to function by a unique mechanism at the level of ODC messenger RNA expression. Based on our findings, rotenoids can be regarded as promising new chemopreventive or anticancer agents.

From the defatted meal of flaxseed (Linum usitatissimum), a novel phenylpropanoid glucoside, linusitamarin [1], was isolated, along with a number of known compounds. The structure of 1 was determined by spectroscopic analysis.

A new pregnane glycoside, marsdekoiside A was isolated from the stems of Marsdenia koi (Asclepiadaceae) and its structure was elucidated from chemical and spectral data as 12-cinnamoyl-dihydrosarcostin-3-O-methyl-6-deoxy-beta-D-allopyr ano syl-(1----4)-O-beta-D-oleandropyranosyl-(1----4)-O-beta-D-++ +cymar opyranoside.

By random screening test, Marsdenia koi was found to have antifertility activity on SD rat. From MeOH extracts of this plant two steroidal glycosides, marsdekoiside A and B, were isolated, and their structures were elucidated on the basis of spectral evidence and by comparison of the hydrolysis products with the authentic samples. Both are newly identified compounds, and marsdekoiside A has good antifertility activity.

The pentane and CHCl3 fractions of a crude extract of Michelia floribunda exhibited cytotoxic activity when tested in KB and P388 tumor cell cultures. Repeated chromatography led to the isolation of three cytotoxic sesquiterpene lactones (costunolide, parthenolide, and santamarine) and a cytotoxic isoquinoline alkaloid (liriodenine). Inactive sesquiterpene lactones obtained during the course of this study included dihydroparthenolide and two new glucosides of dihydrotamaulipin A and dihydroreynosin (1 and 2). The structures of these new compounds were determined through interpretation of their spectroscopic data including 2D-NMR spectroscopy. Syringin was also isolated from the extract.

Following bioassay-directed fractionation procedures, two 7-oxygenated-8-prenylflavones, (-)-pseudosemiglabrin [1] and (-)-semiglabrin [2], were isolated from an MeOH extract of Tephrosia semiglabra. (-)-Pseudosemiglabrin [1] displayed in vitro inhibitory effects on human platelet aggregation.

From the leaves and stems of Larrea tridentata six new furanoid lignans, compounds 1-6, have been isolated and their structures determined through interpretation of physical and spectroscopic properties. The use of 1D and 2D nOe experiments was of particular importance in assigning the stereochemistry.

Rutaecarpine (1) and dehydroevodiamine (2) were isolated from the unripe fruit of Evodia rutaecarpa. In vitro uterotonic assay on rat uterus showed both alkaloids to be active. Dehydroevodiamine was further shown to be active in the in vivo uterotonic assay using rats as the animal model. Reference evodiamine hydrochloride (3) was also evaluated and found to be devoid of uterontonic activity at the doses administered.

A phytochemical investigation of the leaves and twigs of Helietta parvifolia (Rutaceae) resulted in the isolation of heliparvifoline, a new furoquinoline alkaloid, in addition to the known bases flindersiamine and isoflindersiamine.

A reversed phase HPLC method was developed for the qualitative and quantitative analysis of selected biologically active diterpenoid constituents of methanolic and hot water extracts of some MONTANOA species.

Employing the roots of Iris missouriensis, two known triterpenes, iso-iridogermanal and zeorin, were isolated and identified. As presently reported, they were found to demonstrate cytotoxic activity toward cultured P-388 cells (ED50 = 0.1 and 1.1 microgram/mL, respectively). Additionally, a new triterpene that demonstrated an ED50 of 8.5 micrograms/mL was isolated. On the basis of spectral analysis and chemical correlation with zeorin, the structure was shown to be 6 6 alpha-hydroxy-A'-neogermmacer-22(29)-en-30-oic acid (missourin).

Aristolochic acid (1), a constituent of Aristolochia species, has been used for medicinal purposes since the Graeco-Roman period. Following the observation that the compound was mutagenic and carcinogenic, it was removed from pharmaceutical products. Consistent with previous reports, we have found that 1 serves as a direct-acting mutagen in Salmonella typhimurium strains TA100, TA102, TA1537 and TM677, but was not active in the nitroreductase-deficient strains TA98NR and TA100NR. However, aristolic acid (2), a compound that differs in structure only by the absence of the nitro group, was also found to be a direct-acting mutagen in Salmonella strains TA98, TA100, TA102, TA1537, and TM677, as well as strains TA98NR and TA100NR. Both compounds (1 and 2) were active mutagens when evaluated with cultured Chinese hamster ovary cells. Thus, in contrast to previous suggestions, the nitro group at position 10 is not required to induce a mutagenic response. Also, a series of structural relatives (the methyl esters of 1 and 2 (3 and 4, respectively), aristolochic acid-D (5), aristolactam (6), aristolactam A-II (7), and aristolactam-N-beta-D-glucoside (8)) were evaluated for mutagenic potential with Salmonella typhimurium strain TM677 and found to be inactive. Since compounds 3 and 4 were found to be active mutagens with Salmonella typhimurium strains TA98, TA100, TA102 and TA1537 (sufficient quantities of compounds 5-8 were not available for testing), differential sensitivity of the tester strains unrelated to mutagenic potential is suggested. Further, compounds 1, 2, and 6-8 were evaluated for potential to inhibit growth with cultured KB or P388 cells. P388 cells were substantially more sensitive, and compound 1 was the most active of the materials tested (ED5 = 0.58 microM). Compound 6 also demonstrated appreciable activity (ED50 = 4.2 microM), as did compound 8 (ED50 = 6.0 microM). It therefore appears that phenanthrene-ring substituents, in addition to the nitro group at position 10, serve important roles for biological potential. In considering the carcinogenic event induced by aristolochic acid, these functionalities should also be taken into account.

As part of a continuing effort to provide novel agents of potential value in cancer chemotherapy, 2-hydroxy-3-octadecyl-5-methoxy-1,4-benzoquinone (irisoquin) was identified as a component of Iris missouriensis. This novel species demonstrated cytotoxic activity with cultured KB and P-388 cells (ED50 = 1.8 and 0.03 microgram/mL, respectively). The structure was assigned on the basis of spectral analyses and The structure was assigned on the basis of spectral analyses and confirmed by chemical synthesis. The latter provides a facile method for the production of irisoquin and structural derivatives that may be of value for the examination of structure-activity relationships. A closely related compound, 3-octadecyl-5-methoxy-1,4-benzoquinone (deoxyirisoquin), was also isolated from Iris missouriensis, prepared synthetically, and found to be devoid of cytotoxic activity.

The zoapatle plant, Montanoa tomentosa, has an extensive ethnomedical history of use in fertility regulation. Several fractions and isolates of this plant were evaulated in in vitro and in vivo tests to identify and characterize active constituents which may be responsible for its antifertility effects. The guinea pig was the animal model. Two types of in vitro activity were observed, a uterotonic type effect and an inhibition of spontaneous contractions of the uterine muscle. The in vivo effects appear to correlate with the ability of a fraction/isolate to inhibit the spontaneous contractions in in vitro testing. The non-polar fractions/isolates accounted for the inhibition of spontaneous contraction effects and the in vivo activity. The more polar fractions/isolates accounted for the uterine stimulant activity.

An aqueous ethanol extract of Eupatorium altissimum L. (Compositae) showed confirmed activity in the P-388 lymphocytic leukemia assay in mice, and the chloroform solubles showed both cytotoxic activity in the 9KB carcinoma of the nasopharynx cell culture assay and antitumor activity in the P-388 lymphocytic leukemia assay. Two flavones, eupatorin and 5-hydroxy-3',4',6,7-tetramethoxyflavone, were isolated and identified. Both were devoid of cytotoxic and antitumor activity.