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Toranja 'Burarama', Citrus maxima (Burm.) Merr. (Citrus grandis), is a new citrus discovered in the State of Espirito Santo, Brazil. As several varieties of citrus are known to possess antioxidant and cancer chemopreventive properties, the aim of the study was to evaluate in vitro if this Toranja possess these properties. The antioxidant activity, the potential to induce quinone reductase 1, and the influence on cell viability were measured. ESI(-)FT-ICR MS analysis was also performed and identified flavonoids, coumarins, and fatty acids in the extract. The ethyl acetate and methanolic extracts of the peels presented the highest antioxidant activity in vitro by DPPH (IC50 = 298.3 ± 2.6 μg/ml and 303.8 ± 0.4 μg/ml), ABTS assay (IC50 = 298.2 ± 6.4 μg/ml and 296.4 ± 2.5 μg/ml), and FRAP (IC50 = 234.6 ± 1.8 μg/ml and 398.1 ± 3.8 μg/ml). The ethyl acetate extract of the peel induced quinone reductase 1 activity in Hepa1c1c7 cells, indicating that C. maxima exhibited cancer chemopreventive properties.

Scope Physalis peruviana (Solanaceae) is used for culinary and medicinal purposes. We currently report withanolides, isolated from P. peruviana, inhibit the growth of colon cancer monolayer and spheroid cultures. A detailed mechanistic evaluation was performed with 4β-hydroxywithanolide E (4HWE). Methods and results Treatment of HT-29 cells with low concentrations of 4HWE inhibited growth while enhancing levels of p21(Waf1/Cip1) and reducing levels of several cell cycle-related proteins. Apoptosis was induced at higher concentrations. In addition, 4HWE treatment downregulated the levels of Hsp90 client proteins. Nuclear sirtuin 1 (SIRT1) was increased and histone H3 acetylated at lysine 9 was decreased. An additional consequence of SIRT1 elevation in the nucleus may be inhibition of c-Jun activity. The expression of 21 genes was altered, including downregulation of PTGS2, and this correlated with reduced protein levels of cyclooxygenase-2 (COX-2). Overall, efficacious induction of G0/G1 cell cycle arrest at low concentrations, and induction of apoptosis at higher concentrations are interesting 4HWE-mediated phenomena that are accompanied by a complex array of molecular events. Conclusion Considering the worldwide prevalence of colon cancer, and the unique mode of action mediated by 4HWE, it is reasonable to investigate additional mechanistic details and the potential utility of this compound.

Two new heteroleptic palladium(II) complexes have been synthesized by reacting equimolar quantities of palladium(II) chloride, sodium 4-(2-methoxyphenyl)piperazine-1-carbodithioate and diphenyl-p-tolylphosphine (1) or tri-p-tolylphosphine (2). Complexes 1 and 2 have been characterized using elemental analysis, Fourier transform infrared spectroscopy, multinuclear NMR (1H, 13C and 31P) spectroscopy and single-crystal X-ray analysis. The latter technique confirms a pseudo square-planar geometry in which two adjacent positions are occupied by bidentate dithiocarbamate while chloro and substituted triphenylphosphine are present at the remaining two positions. The anticancer activity of both complexes against five different cancer cell lines (LU – human lung carcinoma, established at UIC, Department of Surgical Oncology; MCF7 – human breast adenocarcinoma, ATCC number HTB-22™; MDA-MB-231 – human breast adenocarcinoma, ATCC number HTB-26™; Hepa-1c1c7 – mouse liver hepatoma, ATCC number CRL-2026™; PC-3 – human prostate adenocarcinoma, ATCC number CRL-1435™) was determined by MTT assay, revealing 2 has higher activity than 1. A drug–calf thymus DNA binding study with UV–visible spectroscopy reveals a higher DNA binding affinity of 2 (3.511 × 104 M−1) than 1 (4.213 × 103 M−1). Density functional theory studies confirm the relatively more stable nature of 2 than 1. Copyright © 2016 John Wiley & Sons, Ltd.

Traditional medicinal plants are often used for both the prevention and the treatment of local diseases. Taking into consideration the medicinal importance of Hedera nepalensis within local Pakistani traditions, the present study was undertaken to analyze the in vitro cancer chemopreventive and cytotoxic properties of the plant. The in vitro cancer chemopreventive testing was performed using nitrite assay, NFκB assay, aromatase assay, and quinone reductase 1 (QR1) assay. The cytotoxic potential was evaluated on three cancer-cell lines: MCF-7, MDA-MB-231, and HeLa using sulforhodamine B (SRB) assay. The results of cancer chemopreventive assays show that n-hexane and ethyl acetate fractions of tested plant have promising cancer chemopreventive potential. Lupeol isolated from n-hexane as well as ethyl acetate fraction showed lowest IC50 (0.20 ± 1.9 μM) in NFκB assay. Crude extract and its fractions inhibited the growth of three cancer cell lines by more than 60%, IC50 value of lupeol varied from 2.32 to 10.2 μM. HPLC-DAD-based quantification of lupeol in different plant tissues demonstrated that leaves of H. nepalensis are a rich source of lupeol (0.196 mg/100 mg dry weight). Our data have shown that H. nepalensis harbors cancer chemopreventive and cytotoxic agents.

Although resveratrol can modulate multiple stages of carcinogenesis, by most common standards it is not a good drug candidate. Resveratrol lacks potency, high efficacy, and target specificity; it is rapidly metabolized and serum concentrations are low. Using resveratrol as a scaffold, we produced over 100 derivatives, some of which have target specificity in the nanomolar range. Aromatase inhibition was enhanced over 6000-fold by using 1,3-thiazole as the central ring of resveratrol. Optimizing the substitution pattern of the two phenyl rings and the central heterocyclic linker led to selective QR1 induction with a CD value of 87 nM. Several derivatives have been selected for evaluation of synergistic effects. Preliminary results with pairs of compounds are promising and further experiments, in a constant multidrug manner, will allow us to create polygonograms for larger combinations of derivatives. The objective is to develop a highly efficacious cocktail of derivatives based on the structure of resveratrol.

Several lactone- and lactam-based neoflavonoids and tetrahydroquinolones were synthesized and evaluated for cancer chemopreventive studies using cell and molecular target-based in vitro bioassays, namely NFκB, aromatase, and quinone reductase 1. These analogs blocked TNF-α-induced NFκB activation in a dose-dependent manner with IC₅₀ values in the range of 0.11-3.2 μM. In addition, compound 8 inhibited aromatase activity with an IC₅₀ value of 12.12 μM, and compound 10 affected quinone reductase 1 induction (IR, 3.6; CD, 19.57 μM). Neoflavonoids 8 and 10 exhibiting good results can further be optimized for improved therapeutic profiles. However, investigations into the actions of neoflavonoids and tetrahydroquinolones, especially those related to the NFκB signaling pathway, aromatase inhibition, induction of quinone reductase 1 expression, and in vivo studies could provide new insights into the cancer chemopreventive ability of these molecules.

Cancer chemoprevention entails the ingestion of dietary or pharmaceutical agents that can prevent, delay, or reverse the process of carcinogenesis. With support provided by the National Cancer Institute, we have been actively engaged in the systematic discovery and characterization of natural chemopreventive agents. The typical approach involves identifying active crude substances such as extracts derived from terrestrial plants or marine organisms, utilizing in vitro bioassay systems, followed by the isolation of pure active components. As part of this project, an extract obtained from a nonedible Peruvian legume, Cassia quinquangulata Rich. (Leguminosae), was evaluated and found to be active as an inhibitor of cyclooxygenase. The active component was identified as resveratrol. Surprisingly broad spectrum activity was observed, indicative of potential to inhibit carcinogenesis at the stages of initiation, promotion, and progression. This discovery has led to many additional research efforts. There are now around 3,500 papers concerning some aspect of resveratrol action, yet the molecule is unusually promiscuous and specific mechanisms remain elusive. Considering the structural simplicity of this stilbene, the intensity of interest is phenomenal.

The transcription factor nuclear factor kappa B (NFkappaB) is found in nearly all animal cell types. It is involved in cellular responses to stimuli such as stress, cytokines, free radicals, ultraviolet irradiation, oxidized LDL and microbial antigens, and has been shown to regulate the expression of a number of genes including bcl-2, bcl-xl, cIAP, suvivin, TRAF, COX-2, MMP-9, iNOS and cell cycle-regulatory components. Many carcinogens, inflammatory agents and tumor promoters have been shown to activate NFkappaB, and resulting tumors demonstrate misregulated NFkappaB. Incorrect regulation of NFkappaB has been linked to inflammatory and autoimmune diseases, septic shock, viral infection and improper immune development. Aberrant regulation of NFkappaB is involved in cancer development and progression as well as in drug resistance. Inhibitors of NFkappaB mediate effects potentially leading to antitumor responses or greater sensitivity to the action of antitumor agents. Tools have been developed for the rapid assessment of NFkappaB activity, so in concert with a better understanding of NFkappaB activation mechanisms, many agents capable of suppressing NFkappaB activation have been identified. The present article focuses on the functions of NFkappaB, its role in human cancer and the therapeutic potential and benefit of targeting NFkappaB by natural products in cancer chemoprevention.

Cancer is a leading cause of death worldwide. Cancer chemoprevention is one of the promising strategies to decrease its incidence and both plant extracts and natural products may constitute sources of new chemoprevention agents. Some Brazilian species popularly used to treat inflammatory conditions were selected for evaluation for cancer chemoprevention. A total of 32 extracts/fractions from Hancornia speciosa, Davilla elliptica, Jacaranda caroba, Mansoa hirsuta, Remija ferrugina, Solanum paniculatum and Xyris pterygoblephara, along with a mixture of ursolic and oleanolic acids obtained from J. caroba and a dihydroisocoumarin isolated from aerial parts of X. pterygoblephara were tested for their cancer chemoprevention activity [inhibition of 12-O-tetradecanoyl-13-acetate (TPA)-mediated NF-kappaB activation, ornithine decarboxylase (ODC) and cyclooxygenase-1 (COX-1); induction of antioxidant response element (ARE)]. Several extracts/fractions were active in more than one assay and those from H. speciosa, M. hirsuta and J. caroba mediated strong responses with NF-kappaB, COX-1 and ARE, respectively.

Physalis philadelphica Lam, commonly known as a tomatillo, is a staple of the Mesoamerican cuisine. In our laboratory, an ethyl acetate-soluble extract and four withanolides [ixocarpalactone A (IxoA), ixocarpalactone B, philadelphicalactone B, and withaphysacarpin] were isolated. Studies conducted on Hepa-1c1c7 hepatoma cells revealed that withanolides were potent inducers of quinone reductase, suggesting possible cancer chemoprotective activity. Here we evaluated the antiproliferative properties of the withanolides in SW480 human colon cancer cells. IxoA, which is present in the edible part of the tomatillo, was selected for further evaluation. SW480 cells treated with IxoA showed cell cycle arrest in the G2/M phase, up-regulation of hyper-phosphorylated retinoblastoma, and down-regulation of E2F-1 and DP-1. On the basis of flow cytometry analysis, ethidium bromide/acridine orange, and 4',6-diamidino-2-phenylindole staining, it was found that IxoA induces apoptosis in SW480 cells. Moreover, increased concentrations of the pro-apoptotic protein, BIM/BOD, were found by western blot analysis and immunocytochemistry. Morphological examination revealed vacuole formation in cells treated with IxoA, and Oil Red O staining showed that the vacuole content was nonlipid. Furthermore, immunocytochemistry demonstrated increased concentrations of mucin 3 in IxoA-treated SW480 cells. These findings suggest that chemicals present in tomatillos (e.g. IxoA) may have cancer chemopreventive properties.

In an attempt to determine the relationships between the plant profiles (country of collection, taxonomy, plant part) and the compound classes isolated with cytotoxic activity against a panel of human tumor cell lines, the data compiled from a 15-year anticancer drug-discovery project were subjected to an analysis of variance (ANOVA). The results indicate significant trends in cytotoxic activity relative to collection location, taxonomy, plant part, and compound classes isolated. Plant collections were made in tropical forests in six countries, with collections from Ecuador resulting in higher activity than those from Indonesia and Peru. Interestingly, collections from Florida were not statistically different than those from the countries with higher biodiversity. One hundred and forty-five families were represented in the collections, with the Clusiaceae, Elaeocarpaceae, Meliaceae, and Rubiaceae having low ED50 (half maximal effective dose) values. Especially active genera included Aglaia, Casearia, Exostema, Mallotus, and Trichosanthes. Roots and below-ground plant materials were significantly more active than above-ground materials. Cucurbitacins, flavaglines, anthraquinones, fatty acids, tropane alkaloids, lignans, and sesquiterpenoids were significantly more active than xanthones and oligorhamnosides. The results from this study should serve as a guide for future plant collection endeavors for anticancer drug discovery.

Wild lowbush blueberry fruit extract was fractionated using vacuum chromatography and analyzed for chemopreventive potential using bioassays that test the ability of compounds to inhibit the initiation, promotion, and progression stages of carcinogenesis. A fraction containing phytosterols was active against the initiation stage (quinone reductase assay). However, more polar compounds were inhibitors of later stages of carcinogenesis; a fraction containing flavan-3-ols and fractions containing mainly anthocyanins, phenolic acids, flavan-3-ols, and some proanthocyanidin dimers demonstrated activity against the promotion stage (cyclooxygenase and ornithine decarboxylase assays, respectively), and a proanthocyanidin-rich fraction demonstrated antiproliferation activity (inhibition of cancerous murine hepatocyte proliferation is associated with the progression stage). These results indicate that lowbush blueberries contain a range of compounds that have bioactivity against multiple stages of carcinogenesis, and different types of phenolic compounds are active at different stages.

Bioassay-guided fractionation of the antimalarial-active CHCl3 extract of the dried stem of Nauclea orientalis (L.) L. (Rubiaceae) has resulted in the isolation of two novel tetrahydro-beta-carboline monoterpene alkaloid glucosides, naucleaorine (= (16alpha,17beta)-3,14:15,20-tetradehydro-16-ethenyl-17-(beta-D-glucopyranosyloxy)-19alpha-methoxyoxayohimban-21-one; 1) and epimethoxynaucleaorine (2), as well as the known compounds, strictosidine lactam (= (15beta,16alpha,17beta)-19,20-didehydro-16-ethenyl-17-(beta-D-glucopyranosyloxy)oxayohimban-21-one; 3), 3,4,5-trimethoxyphenol (4), 3alpha-hydroxyurs-12-en-28-oic acid methyl ester (5), 3alpha,23-dihydroxyurs-12-en-28-oic acid (6), 3alpha,19alpha,23-trihydroxyurs-12-en-28-oic acid methyl ester (7), and oleanolic acid (8). Compounds 1, 2, 6, and 8 showed moderate in vitro activities against Plasmodium falciparum. Their structures and configurations were elucidated by spectroscopic methods including 1D- and 2D-NMR analyses.

Summary Gedunin from Cedrela odorata (Meliaceae), a potent in vitro antimalarial agent, was investigated for its in vivo efficacy in CD-1 mice infected with Plasmodium berghei. When orally administered at 50 mg kg -1 day -1 for 4 days, gedunin was able to suppress the parasitaemia level by 44%. However, no clear dose-response effects were observed in the 0-100 mg kg -1 day -1 dose range. Preliminary pharmacokinetics in Sprague-Dawley rats showed poor absorption. However, a binary treatment of 50 mg kg -1 day -1 gedunin with 25 mg kg -1 day -1 dillapiol, a cytochrome P450 inhibitor, increased parasitaemia clearance in mice to 75%. A clear dose-response was observed in the 0-50 mg kg -1 day -1 gedunin dose range when administration was combined with 25 mg kg -1 day -1 dillapiol. In addition, 7- methoxygedunin, a semi-synthetic derivative which is more stable to degradation than gedunin, suppressed the level in mice by 67% at 50 mg kg -1 day -1 . When administered at this dose in combination with 25 mg kg -1 day -1 dillapiol, clearance increased to 80%. These results demonstrate the potential efficacy of antimalarial drugs and phytomedicines based on gedunin and the value of the combination therapy.

A lichen depsidone derivative, fumarprotocetraric acid, was obtained as the major constituent of Cladonia furcata (Huds.) Schrad. collected in Iceland. In the 1H NMR spectrum (in DMSO-d6) of this compound, the trans double bond proton signals of the fumaryl unit appeared as overlapping singlets. To confirm the presence of a trans double bond in the molecule, fumarprotocetraric acid was treated with triethylamine. The 1H NMR spectrum of the resultant triethylamine salt displayed a pair of clear doublets with coupling constants of 15.7 Hz, which are characteristic of a trans double bond. Unambiguous 1H and 13C NMR spectral data for both fumarprotocetraric acid and its triethylamine salt were assigned for the first time based on the interpretation of their 2D NMR (HMQC, HMBC and NOESY) spectra. Copyright © 2003 John Wiley & Sons, Ltd.

Deguelin [(7aS,BaS)-13,13a-dihydro-9,10-dimethoxy-3,3-dimethyl-3H-Bis[1]benzopyrano[3,4-b:6',5'-e]pyran-7(7aH)-one], a naturally occurring rotenone, has shown chemopreventive efficacy in several in vivo and in vitro models. In this report, the effectiveness of deguelin at inhibiting the development of AOM-induced colonic ACF was investigated in CF-1 mice. Loss of hex activity was assessed as a second biomarker. In an initial experiment, animals were given s.c. injections of AOM (10 mg/kg body weight) once a week for 2 weeks to induce ACF. Deguelin and vehicle (corn oil) were administered i.g. 7 days a week. Treatment was initiated 2 weeks prior to the first dose of carcinogen and continued for the duration of the study. The mean number of ACF for the control group was 29.0 +/- 4.3, whereas the mean numbers of ACF in the deguelin groups were 24.8 +/- 2.7, 7.2 +/- 1.5 and 4.6 +/- 1.4 at doses of 2.5, 5.0 and 10.0 mg/kg body weight, respectively. In a similar manner, treatment with deguelin significantly (p < 0.001) suppressed the appearance of hex(-) crypts in a dose-dependent manner. In a second study, the ability of deguelin to block the initiation and promotion stages of colon carcinogenesis was investigated. Greatest inhibition was observed when deguelin was administered during the promotional stage (73.3%, p < 0.001). These results demonstrate that deguelin is an efficacious chemopreventive agent against colon carcinogenesis.

trans-Resveratrol, a phenolic phytoalexin occurring in grapes, wine, peanuts, and cranberries, has been reported to both have anticarcinogenic, antioxidative, phytoestrogenic, and cardioprotective activities, and to be a weak inhibitor of cytochrome P450 (CYP)3A4, which might have significance for drug-drug interactions. Since trans-resveratrol is rapidly converted in vivo to primarily trans-resveratrol-3-sulfate, a rapid, selective, and sensitive method using liquid chromatography/tandem mass spectrometry (LC/MS/MS) was developed to investigate human cytochrome P450 inhibition by trans-resveratrol-3-sulfate. Effects of trans-resveratrol and trans-resveratrol-3-sulfate on the metabolism of selective cytochrome P450 substrates (CYP1A2/ethoxyresorufin, CYP2C9/diclofenac, CYP2C19/(S)-mephenytoin, CYP2D6/bufuralol, CYP3A4/testosterone) were monitored using cDNA-expressed human recombinant isozymes. For method validation, LC/MS/MS was used to measure the inhibition of various cytochrome P450 isozymes by different concentrations (0-50 microM) of known selective inhibitors. IC(50) values of 3.2, 1.4, 8.9, 0.2, and 0.3 microM were obtained for the standard isozyme inhibitors CYP1A2/furafylline, CYP2C9/sulfaphenazole, CYP2C19/tranylcypromine, CYP2D6/quinidine, and CYP3A4/ketoconazole, respectively, which were in good agreement with literature values. trans-Resveratrol showed IC(50) values of 11.6 microM for CYP2C19 and 1.1 microM for CYP3A4, but the IC(50) values exceeded 50 microM for all the other CYP isozymes, which indicated no inhibition. No enzyme inhibition was observed for trans-resveratrol-3-sulfate. Our results indicate that trans-resveratrol is a marginal inhibitor of CYP3A4 and a weak inhibitor of CYP2C19, but its major metabolite trans-resveratrol-3-sulfate is not an inhibitor of any of the cytochrome P450 isozymes investigated.

Activity-guided fractionation of a chloroform-soluble extract of Lonchocarpus aff. fluvialis stem bark using a human epidermoid (KB) tumour cell line as a monitor afforded five rotenoids, one pterocarpan, one chalcone, three flavanones, one flavone and one triterpenoid. All of the compounds isolated proved to be of previously known structure. Among them, the rotenoids (-)-sumatrol and (+/-)-villosinol, the dibenzoylmethane derivative (+)-3,4-methylenedioxy-2'-methoxy-[2",3":4',3']-furanodibenzoylmethane, and the flavanones (-)-isoglabrachromene and (-)-candidone have been shown to exhibit significant cytotoxic activity against human cancer cells for the first time. This is the first report of the chemical constituents of this species, and the profile of compounds obtained was in accordance with the established chemosystematic patterns of species in the tribe Tephrosieae (Leguminosae, Papilionoideae).

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.

Betulinic acid is a naturally occurring pentacyclic triterpenoid. Betulinic acid has recently been selected by the National Cancer Institute for addition into the RAID (Rapid Access to Intervention in Development) programme. The agent exhibits potential anti-tumour activity and functions in this regard via apoptosis. The objective of the present study was to determine the pharmacokinetics of betulinic acid in CD-1 mice. Serum samples were obtained at designed times after a single 250 or 500 mg/kg intraperitoneal (IP) dose of betulinic acid. Tissue samples (skin, heart, liver, spleen, kidney, lung, brain, colon, caecum, ovary, uterus, thymus, lymph node, bladder, perirenal fat, mammary gland and small intestine) were collected after betulinic acid administration (500 mg/kg). Betulinic acid was extracted with methylene chloride and quantitatively analysed by HPLC/MS. Oleanolic acid and madecassic acid were used as internal standards. Pharmacokinetic parameters were calculated using the WinNonlin pharmacokinetic software package. A two-compartment, first-order model was selected for pharmacokinetic modelling. The results showed that after IP 250 and 500 mg/kg betulinic acid, the serum concentrations reached peaks at 0.15 and 0.23 h, respectively. The 250 and 500 mg/kg above betulinic acid IP doses were found to have elimination half-lives of 11.5 and 11.8 h and total clearances of 13.6 and 13.5 L/kg/h, respectively. The pharmacokinetic parameters observed for IP betulinic acid 500 mg/kg in the skin of mice were as follows: k(a) (h(-1)) 0.257, k(10) (h(-1)) 0.234, t(1/2(alpha)) (h) 2.63, t(1/2(beta)) (h) 20.2, V (L/kg) 0.61, AUC (microg/h/mL) 3504, T(max) (h) 3.90 and C(max) (microg/mL) 300.9. The distribution of betulinic acid in tissues at 24 h post-IP administration in a descending order was as follows: perirenal fat, ovary, spleen, mammary gland, uterus, bladder, lymph node, liver, small intestine, caecum, lung, thymus, colon, kidney, skin, heart and brain.

Two alkenyl catechols were rapidly identified as probable cytotoxic constituents from the petroleum ether extract of Semecarpus anacardium using HPLC–electrospray/MS analysis linked to bioassay. These compounds were then isolated from the same extract using semi-prep HPLC and were proven to be 1,2-dihydroxy-3-pentadeca-7′,10′-dienylbenzene and 1,2-dihydroxy-3-pentadec-8′-enylbenzene, respectively, by comparison with physico-chemical data in the literature. Both isolates showed cytotoxic activities against a panel of human cancer cell lines. Copyright © 1999 John Wiley & Sons, Ltd.

In the search for new potently sweet compounds from plants, the rapid identification and quantification of free sugars and polyols in a crude plant extract is important for dereplication purposes, wherein compounds of known structure or biological activity are removed from further consideration. Accordingly, plants found to have high levels of free sugars and polyols are regarded as lower priority leads when screening for novel natural sweeteners. In the present study, gas chromatography–mass spectrometry was used to examine the sugar/polyol content of six sweet-tasting species, comprised of the pericarp of Dialium indum L. (Leguminosae), the stem of Drypetes floribunda Hutchinson (Euphorbiaceae), the fruit of Hymenaea oblongifolia Huber var. palustris (Ducke) Lee and Langenheim (Leguminosae), the rhizomes of Imperata cylindrica (L.) Beauvois (Gramineae), the fruit of Manilkara zapota (L.) van Royen (Sapotaceae), and the pericarp of Sapindus rarak DC. (Sapindaceae). The total yields of sugars/polyols in these plant parts were 1.9, 6.1, 7.8, 4.5, 10.8 and 2.9% w/w, respectively. Several uncommon polyols were identified, including bornesitol in D. floribunda and quebrachitol in S. rarak. It is likely that the sweet taste of the plants containing more than 5% of sugars/polyols is imparted as a result of the high free sugar and/or polyol content. Owing to its low level of free sugars, S. rarak pericarp was chosen for further study, and the known sesquiterpene glycoside mukurozioside IIb (1) was isolated in high yield (6.3% w/w) as a sweet-tasting constituent. Preliminary evaluations, comprised of mouse acute toxicity and bacterial mutagenesis determinations, indicated the safety of 1. The compound was subsequently rated by a human taste panel as having about the same sweetness potency as sucrose. © 1997 by John Wiley & Sons, Ltd.

A library of 2-phenylindole derivatives is synthesized by one-pot palladium-catalyzed Sonogashira-type alkynylation reactions and base-assisted cycloaddition reactions.

Three protoberberine alkaloids, berberine chloride, berberrubine chloride and thalifendine chloride were isolated from the roots of Fibraurea chloroleuca Miers, and found to show significant cytotoxic activity with one or more human cancer cell-lines and cultured P-388 cells. A phytoecdysteroid, 20-hydroxyecdysone, was also isolated as a noncytotoxic compound. Based on comparison with previous investigations, different collections of F. chloroleuca roots appear to exhibit considerable variation in their alkaloidal profiles.

Three xanthone derivatives, gambogic acid, isogambogic acid and isomorellinol, were isolated from the dried latex of Garcinia hanburyi. Two of them, isogambogic acid and isomorellinol, are new. Determinations of the structures and stereochemistry were achieved independently by a series of NMR experiments including COSY, ROESY, HMQC, HMBC and selective INEPT. Isolation of isomorellinol from G. hanburyi provided important chemical evidence to link this species to G. morella. The presence of gambogic acid and isogambogic acid, however, demonstrated the difference between these two species. Cytotoxic evaluation of these isolates revealed that all three were active against KB and drug-resistant KB-V1 cell lines.

Plants continue to provide key leading structures and therapeutic agents for a myriad of debilitating diseases. In this presentation, we will discuss the strategies being used, the structure elucidation and spectroscopic assignment of several classes of compounds with cytotoxic, anti-HIV and antimalarial activity, including xanthones and bisbenzylisoquinoline and Amaryllidaceae alkaloids, and we will provide some details of the bioassay systems being used. The application of various one- and two-dimensional nmr techniques to address opportunities for spectroscopic assignment will be described. Since mankind first displayed susceptibility to disease and injury, the discovery of medicinal agents from renewable resources has been an essential and intrinsic aspect of all evolving cultures. Terrestrial plants are an especially viable opportunity for the discovery of biologically active natural products which may serve as commercially significant entities in their own right, or which may provide lead frameworks for the investigation of modified derivatives possessing enhanced activity and/or reduced toxicity. Three diseases of current concern on a global basis are cancer, AIDS and malaria, and there is a need for continued efforts to discover new template molecules from natural sources for each of these afflictions. This paper describes some of our recent strategies and efforts in each of these therapeutic areas. Anticancer Compounds In the discovery of compounds active against the myriad of diseases known collectively as cancer, two plants, the periwinkle, Catlaaraiitliris roseus, and the May apple, Podopliyllum peltatrim, have provided important compounds (vincaleukoblastine and leurocristine in the former case and podophyllotoxin in the latter), which have stimulated research towards more active and less toxic derivatives. In the 1960s and 1970s, the search for anticancer agents continued through the use of one or two cytotoxicity assays coupled with an iia vivo murine lymphocytic model. As cancer chemotherapy and cell biology evolved, it was found that a number of tumor types were resistant to chemotherapy and agents could be evaluated for their potential to intercede in discrete biological events in tumorigenesis or cell proliferation. Camptothecin was disclosed as an anticancer compound in 1966 and taxol in 1971 by Wall and Wani and their collaborators. But it was their unique mechanisms of action, namely, inhibition of topoisomerase I and of microtubule proliferation, respectively, which rekindled the interest in these compounds after early disappointments in clinical trials. Currently, these compounds and their derivatives are generating substantial chemical, biological, clinical, and even political interest (1). Several years ago we established a battery of human cancer cell lines through which we could examine the possibility that certain agents might display selective cytotoxicity. While we have seen numerous instances of differential cytotoxicity, selective activity (i.e. 100-fold activity difference) has not been observed. We have however, adopted a cadre of human cancer cell lines (breast, lung, hormone-dependent breast, hormone-dependent prostate and colon) for routine testing of extracts and compounds. In addition, a number of mechanism-based assays, topoisomerase I, tubulin binding, protein kinase C inhibition, etc. have been established.

From Hymenocallis littoralis, one new alkaloid, named littoraline, together with 13 known lycorine alkaloids and one lignan, were isolated. The structure and NMR assignments of this new alkaloid were determined by 1D and 2D NMR techniques. Littoraline showed inhibitory activity of HIV reverse transcriptase, and lycorine and haemanthamine showed potent in vitro cytotoxicity.

The anti-HIV agent (±)-calanolide A (1) has been synthesized in a five-step approach starting with phloroglucinol [→ 5 → 6 → 11 → 18 → (±)-1], which includes Pechmann reaction, Friedel−Crafts acylation, chromenylation with 4,4-dimethoxy-2-methylbutan-2-ol, cyclization, and Luche reduction. Cyclization of chromene 11 to chromanone 18 was achieved by employing either acetaldehyde diethyl acetal or paraldehyde in the presence of trifluoroacetic acid and pyridine or PPTS. Luche reduction of chromanone 18 at lower temperature preferably yielded (±)-1. Reduction of chromone 12, synthesized by Kostanecki−Robinson reaction from chromene 11, failed to afford (±)-1. The synthetic (±)-1 has been chromatographically resolved into its optically active forms, (+)- and (−)-1. The anti-HIV activities for synthetic (±)-1, as well as resultant (+)- and (−)-1, have been determined. Only (+)-1 accounted for anti-HIV activity, which was similar to the data reported for the natural product, and (−)-1 was inactive.

From the leaves of Monotes engleri, five prenylated flavanones were isolated as constituents that displayed cytotoxic activity against several human cancer cell lines. Three of these substances are novel, namely, 6-(1,1-dimethylallyl)naringenin, 6-(1,1-dimethylallyl)eriodictyol and 3′-O-methyl-6-(1,1-dimethylallyl)eriodictyol, with the other two active substances being the known flavanones, 6,8-diprenyleriodictyol and hiravanone. Additionally, two novel, but non-cytotoxic, biogenetically related flavanones were isolated, 6-[(2RS)-hydroxy-3-methyl-3-butenyl]-8-prenyleriodictyol and 5,4′-dihydroxy-4″,4″-dimethyl-5″-methyl-5″H-dihydrofurano[2″,3″:6,7]flavanone. The structures of the new compounds were determined by spectral analysis 1D- and 2D-NMR experiments.

Betulinic acid has been modified at C-3, C-20, and C-28 positions and the toxicity of the derivatives has been evaluated against cultured human melanoma (MEL-2) and human epidermoid carcinoma of the mouth (KB) cell lines. This preliminary investigation demonstrates that simple modifications of the parent structure of betulinic acid can produce potentially important derivatives, which may be developed as antitumor drugs.

Vatdiospyroidol (1), a novel cytotoxic resveratrol tetramer, was isolated from the stems of Vatica diospyroides Sym. (Dipterocarpaceae) by bioassay-guided fractionation monitored with a human oral epidermoid carcinoma (KB) cell line. Another novel resveratrol tetramer, vaticaphenol A (2), was obtained as a noncytotoxic constituent, along with the known compounds, bergenin, betulin, betulinic acid, mangiferonic acid, and (E)-resveratrol 3-O-β-d-glucopyranoside. The structures of compounds 1 and 2 were elucidated by spectral analysis, including 1D and 2D NMR experiments, and by molecular modeling.

(+)-(20S)-20-(Dimethylamino)-3α-(methylbenzoylamino)-11-methylene-5α-pregnane (1), a steroidal alkaloid with a new substitution pattern, was isolated from the entire plant of Pachysandra procumbens, together with four other new steroidal alkaloids, (+)-(20S)-3-(benzoylamino)-20-(dimethylamino)-5α-pregn-2-en-4β-ol (2), (+)-(20S)-20-(dimethylamino)-16α-hydroxy-3β-(3′α-isopropyl)-lactam-5α-pregn-4-one (3), (+)-(20S)-20-(dimethylamino)-3α-(methylbenzoylamino)-5α-pregn-12β-yl acetate (4), and (+)-(20S)-20-(dimethylamino)-3α-(methylsenecioylamino)-5α-pregn-12β-ol (5), as well as two known compounds, (+)-pachysamine H (6) and (+)-pachysandrine B (7). The structures of the new compounds were determined by spectroscopic methods. Compounds 1–7 were evaluated for their potential cancer chemopreventive properties using an in vitro estrone sulfatase assay.

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 (IC50 43.7 and 4.7 μM, respectively), while compound 3 was active in 1,1-diphenyl-2-picrylhydrazyl free-radical and cytochrome c reduction antioxidant assays (IC50 29.1 and 16.3 μM, 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.

Four novel stilbenes ( 1 – 4 ) were isolated from the root bark of Ekebergia benguelensis . The structures were established by spectroscopic methods including 2D NMR analysis. The structure of 1 , a new stilbene–coumarin hybrid representing a novel skeleton, was confirmed by single X-ray analysis. Compounds 1 – 4 were evaluated against a panel of human cancer cell lines.

Four novel 29-nor-3,4-seco-cycloartanes, (6S)-hydroxy-25-methoxy-29-nor-3,4-seco-cycloart-4(30),23-dien-3-oic acid methyl ester (1), (6S,25)-dihydroxy-29-nor-3,4-seco-cycloart-4(30),23-dien-3-oic acid methyl ester (2), (6S)-hydroxy-24-oxo-29-nor-3,4-seco-cycloart-4(30),25-dien-3-oic acid methyl ester (3), and (6S)-hydroxy-(24ξ)-hydroperoxy-29-nor-3,4-seco-cycloart-4(30),25-dien-3-oic acid methyl ester (4), along with a semi-synthetic acetylated derivative, (6S,24ξ)-diacetyloxy-29-nor-3,4-seco-cycloart-4(30),25-dien-3-oic acid methyl ester (5), were isolated and characterized from the aerial parts of Antirhea acutata (DC.) Urb. (Rubiaceae). Their structures and absolute stereochemistry were elucidated by spectroscopic and chemical methods. Compounds 1–5 are based on the unprecedented 29-nor-3,4-seco-cycloartane skeleton. Compound 4 showed moderate inhibitory activities in cyclooxygenase-1 and -2 assays (IC50 45.7 and 18.4 μM, respectively), while the other four isolates were inactive.

We report herein the first isolation of a novel structural type sesquiterpene designated as ‘litseane’ from the twigs and leaves of Litsea verticillata Hance (Lauraceae). The isolate (litseaverticillol A, 1 ) was obtained as a racemate through bioassay-guided fractionation and found to inhibit the replication of human immunodeficiency virus (HIV) type 1 with an IC 50 value of 5.0 μg/mL (21.4 μM) and a selectivity index of 2.6. Spectroscopic data and a potential biosynthetic pathway are given.

A new oleanolic acid-derived saponin, polyacetyleneginsenoside-Ro ( 1 ), was isolated along with the known ginsenosides-Ro methyl ester ( 2 ), -Rf, -Rg1, -Rg2, and gingerglycolipid B from the roots of Panax ginseng C. A. Meyer. The new saponin was identified as a ginsenoside-Ro derivative containing a polyacetylene side chain by spectroscopic means including 1D and 2D NMR, and was found to inhibit the replication of human immunodeficiency virus type 1 (HIV-1) with an IC50 value of 13.4 μg/mL (11.1 μM).

A C29-triterpene, beccaridiol (1), a dihydrochalcone natural product, 2',4'-dihydroxy-3-(4-methoxyphenyl)-propiophenone (2), as well as three known compounds, 4'-hydroxy-1',2'-dihydro-beta-ionone, 4'-O-methyldavidigenin (3), and ursolic acid, have been isolated from an EtOAc-soluble extract of the leaves of Diplectria beccariana. Beccaridiol (1) was characterized as an ursane-type 28-nortriterpene possessing an unusual aromatic E-ring by spectroscopic data interpretation. The relative configuration of this unusual isolate was established by analyzing the observed NOESY NMR correlations, and the absolute stereochemistry of 1 was then determined based on the circular dichroism (CD) spectrum of its 2,3-di-p-bromobenzoate (1b) derivative. All isolates were evaluated for their potential cancer chemopreventive properties utilizing a cell culture assay to determine quinone reductase induction.

Bioassay directed-fractionation led to the identification of litseaverticillols A–H ( 1 – 8 ) from the leaves and twigs of Litsea verticillata Hance. These new sesquiterpenes possess a unique skeleton that was recently designated as ‘litseane’. The structures of these compounds were determined by spectroscopic means including 1D and 2D NMR data. Structural configurations were determined by ROESY experiments. Mosher ester reactions and optical rotation measurements established the sesquiterpenes 1 – 8 as racemates. Isolates 1 – 8 inhibited HIV-1 replication in HOG.R5 cells with IC 50 values ranging from 2 to 15 μg/ml (8–58 μM) while affecting the growth of HOG.R5 at concentrations 2–3-fold higher. Based on this data, structure–activity relationships can be discerned, suggesting compounds of this class are good candidates for analog production.

Two new cardenolides, (−)-14-hydroxy-3β-(3- O -methyl-6-deoxy-α- l -rhamnosyl)-11α,12α-epoxy-(5β,14β,17βH)-card-20(22)-enolide ( 1 ), (−)-14-hydroxy-3β-(3- O -methyl-6-deoxy-α- l -glucopyranosyl)-11α,12α-epoxy-(5β,14β,17βH)-card-20(22)-enolide ( 2 ), and a known cardenolide, (−)-17β-neriifolin ( 3 ), were isolated from the roots of Cerbera manghas as antiproliferative and antiestrogenic principles when evaluated against a human colon cancer cell line (Col2) and the Ishikawa cell line, respectively. Two known lignans, (−)-olivil ( 4 ) and (−)-cycloolivil ( 5 ), were also isolated but were inactive in the assay systems used.

Three new anthracene derivatives, rubiasins A–C (1–3), were isolated from the combined roots and stems of Rubia cordifolia, and their structures were elucidated by spectroscopic analysis. Their absolute configurations were determined by Mosher ester methodology. A known compound, mollugin (4), was obtained as an active antiproliferative principle by bioassay-monitored fractionation using a human colon cancer (Col2) cell line.