Pharmacokinetics of 23-epi-26-deoxyactein in women after oral administration of a standardized extract of black cohosh.

Dietary supplements containing black cohosh are alternatives to conventional hormone replacement therapy in menopause. This study investigates the maximum tolerated dose of a 75% ethanol extract of black cohosh and determines the pharmacokinetics of one of its most abundant triterpene glycosides, 23-epi-26-deoxyactein. Single doses of black cohosh extract containing 1.4, 2.8, or 5.6 mg of 23-epi-26-deoxyactein were administered to 15 healthy, menopausal women. Serial blood samples and 24-h urine samples were obtained; blood chemistry, hormonal levels, and 23-epi-26-deoxyactein levels were determined. No acute toxicity or estrogenic hormone effects were observed. Pharmacokinetic analyses of 23-epi-26-deoxyactein in sera indicated that the maximum concentration and area under the curve increased proportionately with dosage, and that the half-life was ~2 h for all dosages. Less than 0.01% of the 23-epi-26-deoxyactein was recovered in urine 24 h after administration. No phase I or phase II metabolites were observed either in clinical specimens or in vitro.

Activation of the mu-opiate receptor by Vitex agnus-castus methanol extracts: implication for its use in PMS.

The dried ripe fruit of Vitex agnus-castus L. (VAC) is widely used for the treatment of premenstrual syndrome (PMS). A previous study reported that extracts of VAC showed affinity to opiate receptors; however, functional activity was not determined. We tested two different VAC extracts in receptor binding and functional assays. Our objectives were: (1) to confirm the opiate affinity; (2) to rule out interference by free fatty acids (FFA); (3) to determine the mode of action of VAC at the mu-opiate receptor. Methanol extracts of VAC were prepared either before (VAC-M1) or after (VAC-M2) extraction with petroleum ether to remove fatty acids. Both extracts showed significant affinities to the mu-opiate receptor, as indicated by the concentration-dependent displacement of [3H]DAMGO binding in Chinese hamster ovary (CHO)-human mu-opiate receptor (hMOR) cells. The IC50 values were estimated to be 159.8 microg/ml (VAC-M1) and 69.5 microg/ml (VAC-M2). Since the defatted extract not only retained, but exhibited a higher affinity (p<0.001), it argued against significant interference by fatty acids. In an assay to determine receptor activation, VAC-M1 and VAC-M2 stimulated [35S]GTPgammaS binding by 41 and 61% (p<0.001), respectively. These results suggested for the first time that VAC acted as an agonist at the mu-opiate receptor, supporting its beneficial action in PMS.

The pharmacognosy of Humulus lupulus L. (hops) with an emphasis on estrogenic properties.

As the population ages, there is an ever-increasing need for therapeutic agents that can be used safely and efficaciously to manage symptoms related to postmenopausal estrogen deficiency. Endogenous estrogens, e.g., 17beta-estradiol, of exogenous mammalian origin, e.g., horses, have long been used to manage such symptoms. There are more than 20 different classes of phytochemicals that have demonstrated affinity for human estrogen receptors in vitro. Some studies on exogenous estrogenic substances of botanical origin (phytoestrogens), such as standardized formulations of plant extracts with in vitro and in vivo estrogenic activity from soy (Glycine max Merill.) and red clover (Trifolium pratense L.), suggest clinical efficacy. Few clinical data for phytoestrogens other than isoflavonoids are available. In an exhaustive review of the literature through 2003, only two clinical trials were identified that were designed to evaluate the effect of hops (Humulus lupulus L.) on symptoms related to menopause. Folkloric, chemical, and biological literature relating primarily to the use of hops for their estrogenic activity, and two human clinical trials, are reviewed.

Chemical and biological characterization and clinical evaluation of botanical dietary supplements: a phase I red clover extract as a model.

Botanical dietary supplements, as compared with nutritional supplements or single-component pharmaceutical drugs, are typically less-refined preparations derived from bulk plant material and, as such, require a modified approach to their development, production, and evaluation. An integrated, multidisciplinary team of scientific and clinical investigators is required in order to develop high quality phytomedicines and rigorously evaluate their safety and efficacy. Research on botanicals involves unique challenges as plant source materials frequently vary in chemical content and may contain unwanted pesticides, heavy metals, contaminant plant species, or other adulterants. Ideally, a botanical formulation should be standardized, both chemically and biologically, by a combination of analytical techniques and bioassays. This combination approach provides multiple measures by which reproducible quality and efficacy of botanical supplements may be achieved, and is particularly useful for botanical products for which the active compound(s) have not yet been identified. Safety and toxicity should be evaluated during the supplement development process in both in vitro and in vivo systems. A number of liquid chromatography-mass spectrometry methods can aid in the assessment of purity, bioavailability, toxicity, metabolism, and molecular target profiling of botanical extracts. Clinical investigators must appreciate the complexity of multi-component phytomedicines and adjust trial protocols accordingly. This review highlights practical considerations of value to basic science and clinical investigators engaged in the study of botanical supplements. Lessons and examples are drawn from the authors' experience in designing and developing a red clover (Trifolium pratense L.) standardized extract for evaluation in Phase I and Phase II clinical trials.

Isolation of linoleic acid as an estrogenic compound from the fruits of Vitex agnus-castus L. (chaste-berry).

A methanol extract of chaste-tree berry (Vitex agnus-castus L.) was tested for its ability to displace radiolabeled estradiol from the binding site of estrogen receptors alpha (ERalpha) and beta (ERbeta). The extract at 46 +/- 3 microg/ml displaced 50% of estradiol from ERalpha and 64 +/- 4 microg/ml from ERbeta. Treatment of the ER+ hormone-dependent T47D:A18 breast cancer cell line with the extract induced up-regulation of ERbeta mRNA. Progesterone receptor (PR) mRNA was upregulated in the Ishikawa endometrial cancer cell line. However, chaste-tree berry extract did not induce estrogen-dependent alkaline phosphatase (AP) activity in Ishikawa cells. Bioassay-guided isolation, utilizing ER binding as a monitor, resulted in the isolation of linoleic acid as one possible estrogenic component of the extract. The use of pulsed ultrafiltration liquid chromatography-mass spectrometry, which is an affinity-based screening technique, also identified linoleic acid as an ER ligand based on its selective affinity, molecular weight, and retention time. Linoleic acid also stimulated mRNA ERbeta expression in T47D:A18 cells, PR expression in Ishikawa cells, but not AP activity in Ishikawa cells. These data suggest that linoleic acid from the fruits of Vitex agnus-castus can bind to estrogen receptors and induce certain estrogen inducible genes.

Antimycobacterial evaluation of Peruvian plants.

We present the results of an antimycobacterial screening of 270 Peruvian plant samples representing 216 species from 171 genera in 63 families. Dichloromethane extracts were tested at a concentration of 50 microg/ml for inhibition of Mycobacterium tuberculosis in radiometric culture. Slightly more than half of the samples tested showed inhibition of M. tuberculosis at this concentration.

Evaluation of commercial Ginkgo biloba dietary supplements for the presence of colchicine by high-performance liquid chromatography.

A high-performance liquid chromatographic method with photodiode array detection was developed for the detection of the presence of colchicine in commercial ginkgo products. The method is based on the baseline separation of constituents in ginkgo samples plus reference colchicine. The minimal detectable concentration of colchicine is 1.0 ng on column in the current assay. By analysis of retention time and UV profile of suspect peaks in the sample with those of reference colchicine, none of the nine commercial ginkgo products analyzed contained colchicine.

A preliminary RAPD-PCR analysis of Cimicifuga species and other botanicals used for women's health.

Traditional taxonomic methods of botanical identification that rely primarily on morphological observations cannot be used efficiently when only powdered plant materials are available. Thus, our objectives were to determine if we could apply a molecular approach to: a) produce unique DNA profiles that are characteristic of the species, and b) determine if the geographical area or time of collection influences these DNA profiles. Towards this end, random amplified polymorphic DNA (RAPD) analyses were performed on a number of botanicals currently used for women's health. The test materials included samples from three species each of the genera Cimicifuga (Actaea) and Trifolium, as well as samples of Vitex agnus-castus L., Glycyrrhiza glabra L., Gingko biloba L., Valeriana officinalis L., Angelica sinensis (Oliv.) Diels, Viburnum prunifolium L., Humulus lupulus L., Vaccinium macrocarpon Ait., Panax ginseng C.A. Mey. Cimicifuga racemosa (L.) Nutt. and Trifolium pratense L. are currently under clinical investigation in our basic research laboratories and medical clinic for the relief of post-menopausal symptoms. Characteristic profiles produced with the OPC-15 primer could distinguish the three Cimicifuga species: C. racemosa, C. americana and C. rubifolia. Similar results were obtained with the three Trifolium species: Trifolium pratense L., Trifolium incarnatum L., and Trifolium repens L. Accessions of cultivated T. pratense collected from the same field at different times, produced identical profiles. Accessions of Cimicifuga species collected from different geographical areas produced similar but not identical DNA profiles; however, species-specific DNA fragments were identified. These results demonstrate that RAPD analysis can be applied to distinguish species when only powdered material is available for testing. This methodology can be applied to identify species of commercial value regardless of collection time or geographic area.

Macharistol, a new cytotoxic cinnamylphenol from the stems of Machaerium aristulatum.

A new cinnamylphenol, macharistol (1), along with a known pterocarpan, (+)-medicarpin (2), were isolated as cytotoxic constituents from the stems of Machaerium aristulatum. In addition, a known pterocarpan, (+)-maackiain (3), and a known isoflavone, formononetin (4), were identified as inactive constituents. Compound 1 was evaluated in the in vivo hollow fiber assay with KB, Col-2, and hTERT-RPE1 cells and found to be inactive at the highest dose (25 mg/kg body weight) tested.

Aromatase inhibitors from Broussonetia papyrifera.

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.

Constituents of Eugenia sandwicensis with potential cancer chemopreventive activity.

A triterpenoid, 3beta-cis-p-coumaroyloxy-2alpha,23-dihydroxyolean-12-en-28-oic acid (1), and two natural products, 3beta-trans-p-coumaroyloxy-2alpha,23-dihydroxyolean-12-en-28-oic acid (2) and 23-trans-p-coumaroyloxy-2alpha,3beta-dihydroxyolean-12-en-28-oic acid (3), were isolated from a chloroform-soluble extract of the stems of Eugenia sandwicensis, along with 10 known compounds. Of these compounds, 2 showed significant inhibitory activity (79.2% at 4 microg/ml) in a 7,12-dimethylbenz[a]anthracene-induced mouse mammary organ culture assay system of relevance to cancer chemoprevention. Gallic acid was isolated as an antioxidative constituent of an ethyl acetate-soluble extract of E. sandwicensis stems. Isolates 1-3 were characterized on the basis of spectral and chemical evidence.

Cytotoxic polyacetylenes from the twigs of Ochanostachys amentacea.

Bioassay-guided investigation of the twigs of Ochanostachys amentacea using LNCaP (hormone-dependent human prostate cancer) cells as a monitor led to the isolation of three alkynes, the known (S)-17-hydroxy-9,11,13,15-octadecatetraynoic acid (minquartynoic acid, 1) and two novel analogues, (S)-17,18-dihydroxy-9,11,13,15-octadecatetraynoic acid (2) and (S)-17-hydroxy-15E-octadecen-9,11,13-triynoic acid (3). Compounds 1-3 were tested against a panel of human tumor cell lines and found to be significantly cytotoxic.

Chemical composition and potential health effects of prunes: a functional food?

Prunes are dried plums, fruits of Prunus domestica L., cultivated and propagated since ancient times. Most dried prunes are produced from cultivar d'Agen, especially in California and France, where the cultivar originated. After harvest, prune-making plums are dehydrated in hot air at 85 to 90 degrees C for 18 h, then further processed into prune juice, puree, or other prune products. This extensive literature review summarizes the current knowledge of chemical composition of prunes and their biological effects on human health. Because of their sweet flavor and well-known mild laxative effect, prunes are considered to be an epitome of functional foods, but the understanding of their mode of action is still unclear. Dried prunes contain approximately 6.1 g of dietary fiber per 100 g, while prune juice is devoid of fiber due to filtration before bottling. The laxative action of both prune and prune juice could be explained by their high sorbitol content (14.7 and 6.1 g/100 g, respectively). Prunes are good source of energy in the form of simple sugars, but do not mediate a rapid rise in blood sugar concentration, possibly because of high fiber, fructose, and sorbitol content. Prunes contain large amounts of phenolic compounds (184 mg/100 g), mainly as neochlorogenic and chlorogenic acids, which may aid in the laxative action and delay glucose absorption. Phenolic compounds in prunes had been found to inhibit human LDL oxidation in vitro, and thus might serve as preventive agents against chronic diseases, such as heart disease and cancer. Additionally, high potassium content of prunes (745 mg/100 g) might be beneficial for cardiovascular health. Dried prunes are an important source of boron, which is postulated to play a role in prevention of osteoporosis. A serving of prunes (100 g) fulfills the daily requirement for boron (2 to 3 mg). More research is needed to assess the levels of carotenoids and other phytochemicals present in prunes to ensure correct labeling and accuracy of food composition tables in order to support dietary recommendations or health claims.

The potential of alkaloids in drug discovery.

Alkaloids are an important group of diversely distributed, chemically, biologically and commercially significant natural products. This article suggests why now, with the presently available technology, and the remaining biome available and reasonably accessible, is an opportune moment to consciously focus on the discovery of further alkaloids with pharmacophoric utility.

Evaluation of estrogenic activity of plant extracts for the potential treatment of menopausal symptoms.

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.

Biological screening of rain forest plot trees from Palawan Island (Philippines).

Study plots totaling 0.2 Ha were established in primary forest in the highlands of central Palawan Island, Philippines. Samples of various anatomical parts [typically leaf + twig (If/tw), stem bark (sb), and root (rt)] were collected from all tree species represented within the plots by individuals having a diameter at breast height > or = 10 cm. In all, 211 distinct samples were obtained from 68 tree species, representing 35 families (not including samples from 4 indeterminate species). Methanol extracts of these samples were tested in in vitro antiplasmodial, brine shrimp toxicity, and cytotoxicity assays. The following samples showed an IC50 < or = 10 microg/mL against either chloroquine-sensitive or chloroquine-resistant clones of Plasmodium falciparum: Acronychia laurifolia (sb), Agathis celebica (lf/tw), Aglaia sp. 1 (sb), Aglaia sp. 2 (lf/tw, rt), Bhesa sp. 1 (rt), Cinnamomum griffithii (lf/tw), Croton leiophyllus (rt), Dysoxylum cauliflorum (rt), Garcinia macgregorii (sb), Lithocarpus sp. 1 (rt, sb), Meliosma pinnata ssp. macrophylla (lf/tw, rt), Myristica guatteriifolia (lf/tw), Ochrosia glomerata (rt, sb), Swintonia foxworthyi (lf/tw), Syzygium sp. 1 (rt), Turpinia pomifera (rt), and Xanthophyllum flavescens (sb). Secondly, those samples which displayed > or = 50% immobilization of brine shrimp at 100 microg/mL were: Acronychia laurifolia (lf/tw/fruit, rt, sb), Agathis celebica (lf/tw, sb), Aglaia sp. 1 (lf/tw), Alphonsea sp. 1 (rt), Ardisia iwahigensis (lf/tw), Arthrophyllum ahernianum (lf/tw, rt, sb), Castanopsis cf. evansii (rt), Cinnamomum griffithii (lf/tw, rt), Croton argyratus (lf/tw), C. leiophyllus (lf/tw, rt), Dysoxylum cauliflorum (fruit, lf/tw, rt), Euonymus javanicus (rt), Glochidion sp. 1 (rt), Polyosma sp. 1 (rt), Symplocos polyandra (rt), Timonius gammillii (sb), and Xanthophyllum flavescens (rt). Lastly, samples which exhibited an IC50 < or = 20 microg/mL against one or more of the cancer cell lines employed (LU1, KB, KB-V1, P-388, LNCaP, or ZR-75-1) include: Acronychia laurifolia (lf/tw/fruit, rt, sb), Aglaia sp. 1 (sb), Aglaia sp. 2 (rt), Alphonsea sp. 1 (rt), Ardisia iwahigensis (lf/tw, rt, sb), Astronia cumingiana (sb), Croton argyratus (lf/tw, rt, sb), C. leiophyllus (lf/tw, rt), Dimorphocalyx murina (lf/tw, rt, sb), Lithocarpus caudatifolius (rt, sb), Litsea cf. sibuyanensis (rt), Syzygium cf. attenuatum (rt, sb), S. confertum (sb), Ternstroemia gitingensis (rt), and Ternstroemia sp. 1 (rt, sb).

Bioactive constituents of the roots of Licania intrapetiolaris.

Fractionation of a methanol extract of the roots of Licania intrapetiolaris, as directed by activity against the KB assay, has led to the isolation of two novel clerodane diterpenoids, intrapetacins A (1) and B (2), and the known triterpenoid cucurbitacin B (3). The structures of 1 and 2 were deduced from one- and two-dimensional NMR experiments, including relative stereochemical assignments based on NOESY correlations and COSY coupling constants. Compound 3 was the most potent against the KB assay, but both 1 and 2 displayed moderate cytotoxicity. When evaluated against an antifungal assay using Aspergillus niger, 2 caused a significant zone of inhibition of fungal growth, while 1 was completely inactive. To the best of our knowledge, this is the first report of the isolation of bioactive compounds from the genus Licania.

The value of plants used in traditional medicine for drug discovery.

In this review we describe and discuss several approaches to selecting higher plants as candidates for drug development with the greatest possibility of success. We emphasize the role of information derived from various systems of traditional medicine (ethnomedicine) and its utility for drug discovery purposes. We have identified 122 compounds of defined structure, obtained from only 94 species of plants, that are used globally as drugs and demonstrate that 80% of these have had an ethnomedical use identical or related to the current use of the active elements of the plant. We identify and discuss advantages and disadvantages of using plants as starting points for drug development, specifically those used in traditional medicine.

Modulation of the multidrug-resistance phenotype by new tropane alkaloid aromatic esters from Erythroxylum pervillei.

Nine tropane alkaloid aromatic esters (1-9) were isolated from the roots of Erythroxylum pervillei by following their potential to reverse multidrug-resistance with vinblastine-resistant oral epidermoid carcinoma (KB-V1) cells. All isolates, including seven new structures (3-9), were evaluated against a panel of human cancer cell lines, and it was found that alkaloids 3 and 5-9 showed the greatest activity with KB-V1 cells assessed in the presence of vinblastine, suggesting that these new compounds are potent modulators of P-glycoprotein. Confirmatory results were obtained with human ovarian adenocarcinoma (SKVLB) cells evaluated in the presence of adriamycin and synergistic studies performed with several cell lines from the NCI tumor panel. The structures of the new compounds were determined using spectroscopic techniques. Single-crystal X-ray analysis was performed on the monoester, tropane-3 alpha,6 beta,7 beta-triol 3-phenylacetate (1).

Plants used against cancer - an extension of the work of Jonathan Hartwell.

A survey of plants used ethnomedically against cancer was undertaken, using the NAPRALERT database currently maintained by the Program for Collaborative Research in the Pharmaceutical Sciences, College of Pharmacy, University of Illinois-Chicago. We report over 350 species which are reported to be used against cancer and not cited in the work of Jonathan Hartwell, "Plants used against cancer: a survey", including previously unrepresented genera and families.