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The anthocyanin delphinidin is a natural compound found as water-soluble pigment in coloured fruits and berries. Anthocyanin-rich diets have been proposed to have bone protective effects in humans and mice, but the underlying mechanisms remain unclear. In this study, we used a medaka (Oryzias latipes) osteoporosis model to test the effects of delphinidin on bone cells in vivo. In this model, inducible transgenic expression of receptor-activator of NF-kβ ligand (Rankl) leads to ectopic formation of osteoclasts and excessive bone resorption, similar to the situation in human osteoporosis patients. Using live imaging in medaka bone reporter lines, we show that delphinidin significantly reduces the number of osteoclasts after Rankl induction and protects bone integrity in a dose-dependent manner. Our in vivo findings suggest that delphinidin primarily affects the de novo differentiation of macrophages into osteoclasts rather than the recruitment of macrophages to sites of bone resorption. For already existing osteoclasts, delphinidin treatment affected their morphology, leading to fewer protrusions and a more spherical shape. Apoptosis rates were not increased by delphinidin, suggesting that osteoclast numbers were reduced primarily by impaired differentiation from macrophage progenitors and reduced maintenance of pre-existing osteoclasts. Importantly, and in contrast to previously reported cell culture experiments, no effect of delphinidin on osteoblast differentiation and distribution was observed in medaka in vivo. Our study is the first report on the effects of delphinidin on bone cells in fish embryos, which are a unique model system for compound testing that is suitable for live imaging of bone cell behaviour in vivo.

Introduction: In previous work, we have shown the synergistic effects of combinations of vitamins A, D2, and D3 in reducing the proliferation of HCT-116 colon cancer cells. This combination also induced apoptosis and altered gene expression patterns as determined by transcriptomic profiling. Aims: The aims of this study were to investigate the effects of vitamins A and D in HCT-116 Crl:NU(NCr)-Foxn1nu mouse xenographs. Also, to determine potential mechanisms of action using miRNA-seq and correlated these data with results from RNA-seq.Methods: HCT-116 colon cancer cells were cultured and used for the xenograph study. Crl:NU(NCr)-Male Foxn1nu mice were injected subcutaneously with HCT-116-cells (1 X 106 in 100 µl 50% Matrigel and Sterile PBS), and after 7 days, were treated with vitamin A and D in feed for 21 days. The mice were imaged on day 28, sacrificed and the tumors were excised and measured. RNA was isolated from the HCT-116 cells and tumors, and RNA-seq and miRNA-seq were performed.Results: There was a concentration-dependent reduction in the HCT-116 colon cancer cell viability after treatment with vitamin A and D combinations. In Crl:NU(NCr)-Foxn1nu mice injected with HCT-116 colon tumor cells, treatment with vitamin A (25,000 IU) and vitamin D (4,000 IU) or vitamin A (35,000 IU) and vitamin D (5,000 IU) for 21 days significantly reduced tumor growth by ~38% and ~58%, respectively (p < 0.001). In the HCT-116 cells and excised tumors, treatment upregulated the expression of Bim, Bax, p53, and PTEN, and downregulated AXIN2, ID2 and DDX20 mRNAs, all well-known pro-apoptotic proteins, tumor suppressors, and molecules involved in the epithelial-mesenchymal transition. Ingenuity Pathway Analysis of miRNA-seq paired with RNA-seq showed correlations between miRNAs expression and expression of these genes. For example, miRNA-mRNA correlation pairing showed that treatment downregulated miR-30c-3p and miR-125b-3p which was associated with upregulated TP53. Upregulation of the tumor suppressor PTEN was associated with the downregulation of eleven miRNAs, including oncogenic miR17 and miR21.Conclusion: The results suggest that the combinations of vitamins A and D reduced the HCT-116 tumor burden in mice and altered the expression of miRNAs directly associated with genes in the apoptosis, tumor suppression, and epithelial-mesenchymal transition pathways. The data supports the hypothesis that vitamin A and D combinations impact multiple cancer signaling pathways and thus may be more effective in the treatment and prevention of cancer, as well as reducing cancer metastasis. Keywords: apoptosis, cholcalciferol, colon cancer, DDX20, epithelial-mesenchymal transition, ergocalciferol, p53, RNA-seq, miRNA-seq, PTEN, synergism, transcriptome

Integrated systems biology approaches suggest that combinations of nutrients may be more effective against cancer due to the large number of signaling pathways associated with cancer initiation and promotion. In a previous work, we have reported that combinations of vitamins A (as all trans-retinoic acid, ATRA), D2, and D3 act synergistically to induce apoptosis in colon and gastric cancer cells. In this work, we use whole-genome transcriptomic profiling to detect gene expression changes using RNA-seq to more comprehensively investigate the biological pathways affected by the combination of vitamin D2, D3 and ATRA. HCT-116 colon cancer cells were harvested, RNA was isolated and RNA-seq libraries were prepared using a Universal Plus mRNASeq kit. Sequencing was carried out on NovaSeq 6000. General quality-control metrics were obtained using FastQC and raw reads were aligned to human reference genome hg38 using STAR and BWA MEM. ENSEMBL genes were quantified using FeatureCounts, and differential expression statistics were computed using EdgeR. Specific gene expression was validated using qPCR. Transcriptomic analysis showed that of 26,313 genes analyzed, the expression of 8,402 genes was significantly altered (4030 up-regulated and 4373 down-regulated, FDR+1 and an FDR

Icaritin, a prenylflavonoid isolated from Herbal Epimedii, is well known for its osteoplastic and anti-tumour activities. Here, we examined the cardioprotective actions of icaritin on cardiac H9c2 cells under oxidative stress conditions induced by treatment with tert-butylhydroperoxide (t-BHP). Icaritin at a concentration range between 1 and 4 µM effectively increased cell viability and decreased lactate dehydrogenase (LDH) leakage and reactive oxygen species (ROS) release in t-BHP treated cells, reflecting its ability to reduce t-BHP-induced cell injury. Icaritin also protected cell membrane integrity by preventing the collapse of the mitochondrial membrane potential. In addition, pretreatment of NF-E2-related factor 2 (Nrf2) siRNA and Akt inhibitor abolished the cardioprotective effect of icaritin. Furthermore, Nrf2 siRNA transfection interfered with the effect of icaritin on the inhibition of Ca2+ overload, whereas Akt inhibitor reduced icaritin-induced haemo-oxygenase-1 (HO-1) expression. Collectively, these results suggest that icaritin exerted its cardioprotective effect through the Akt/Nrf2/HO-1 and calcium signalling pathways.

Background: Vitamin D was first hypothesized to play a role in cancer chemoprevention in 1980 when it was observed that there was a higher rate of colon cancer in the Northern USA as compared with populations living in the South. While cholecalciferol (vitamin D3) has been tested in many cancer-cell lines, published results for ergocalciferol (vitamin D2) are lacking for most epithelial cell cancers, and combination studies of both D2 and D3 and vitamin A (retinoic acid) are lacking in general. The goal of the study was to investigate the effects of vitamins D2, D3, and A, alone and in combination on the growth of all gastric and colon cancer cell lines in vitro. Methods: Colon (SW480 and HCT-116) and gastric (AGS and NCI-N87) cell lines were treated with varying concentrations of D2, D3 and all-trans retinoic acid (ATRA) and combinations thereof. Cell viability and cytotoxicity were measured using the CellTiter-Glo® 2.0 assay, while caspase activities and cytotoxicity were determined using the Caspase-Glo® 3/7, Caspase 8, ApoTox-Glo™ Triplex assays. Autophagy was determined using the CYTO-ID® Autophagy Detection Kit 2.0. Gene expression studies were performed using qPCR. Results: Both vitamin D2 and D3 inhibited the growth of all cell lines tested, with IC 50 ranging from 19-56 μM. However, when combined (1:1), the IC 50 for the combination of D2 and D3 was significantly reduced to a range of 5-6.0 μM indicating synergism. ATRA also inhibited the growth of all cell lines tested with an IC 50 range of 2.6 to 5.6 μM. When ATRA was combined with D2 and D3, the IC 50 s were significantly reduced to 0.65 to 1.4 μM, further indicating synergistic effects. In the gastric and colon cancer cell lines, the combination induced apoptosis via caspase 3/7 and 8, increased the Bax/Bcl-2 ratio, while in the SW480 colon cancer line, the combination also induced autophagy. Conclusion: Our data demonstrated that vitamins D2, D3 and ATRA inhibit the proliferation of colon and gastric cancer cells. The combinations of D2+D3 or ATRA+D2+D3 had synergistic effects or additive effects on all cell lines tested, increased Bax expression and the Bax/Bcl-2 ratio, and increased caspase 3/7 and 8 activities to favor apoptosis. In SW480 colon cancer cells the combination also induced autophagy. This data suggest that combinations of vitamins A and D have synergistic effects in colon and gastric cancers, and considering the potential clinical implications, further research is justified. Key words: apoptosis, autophagy cholcalciferol, ergocalciferol, gastric cancer, colon cancer, caspase, synergism

Osteoporosis and osteopenia impact more than 54 million Americans, resulting in significant morbidity and mortality. Alterations in bone remodeling are the hallmarks for osteoporosis, and thus the development of novel treatments that will prevent or treat bone diseases would be clinically significant, and improve the quality of life for these patients. Bone remodeling involves the removal of old bone by osteoclasts and the formation of new bone by osteoblasts. This process is tightly coupled, and is essential for the maintenance of bone strength and integrity. Since the osteoclast is the only cell capable of bone resorption, the development of drugs to treat bone disorders has primarily focused on reducing osteoclast differentiation, maturation, and bone resorption mechanisms, and there are few treatments that actually increase bone formation. Evidence from observational, experimental, and clinical studies demonstrate a positive link between naturally occurring compounds and improved indices of bone health. While many natural extracts and compounds are reported to have beneficial effects on bone, only resveratrol, sulforaphane, specific phenolic acids and anthocyanins, have been shown to both increase bone formation and reduce resorption through their effects on the bone epigenome. Each of these compounds alters specific aspects of the bone epigenome to improve osteoblast differentiation, reduce osteoblast apoptosis, improve bone mineralization, and reduce osteoclast differentiation and function. This review focuses on these specific natural compounds and their epigenetic regulation of bone remodeling.

Disclosures None. SUMMARY Background and Aims: The use of herbs and dietary supplements (HDS) alone or concomitantly with medications can potentially increase the risk of adverse events experienced by the patients. This review aims to evaluate the documented HDS-drug interactions and contraindications. Methods: A structured literature review was conducted on PubMed, EMBASE, Cochrane Library, tertiary literature and Internet. Results: While 85 primary literatures, six books and two web sites were reviewed for a total of 1,491 unique pairs of HDS-drug interactions, 213 HDS entities and 509 medications were involved. HDS products containing St. John’s Wort, magnesium, calcium, iron, ginkgo had the greatest number of documented interactions with medications. Warfarin, insulin, aspirin, digoxin, and ticlopidine had the greatest number of reported interactions with HDS. Medications affecting the central nervous system or cardiovascular system had more documented interactions with HDS. Of the 882 HDS-drug interactions being described its mechanism and severity, 42.3% were due to altered pharmacokinetics and 240 were described as major interactions. Of the 152 identified HDS contraindications, the most frequent involved gastrointestinal (16.4%), neurological (14.5%), and renal ⁄ genitourinary diseases (12.5%). Flaxseed, echinacea, and yohimbe had the largest number of documented contraindications. Conclusions: Although HDSdrug interactions and contraindications primarily concerned a relatively small subset of commonly used medications and HDS entities, this review provides the summary to identify patients, HDS products, and medications that are more susceptible to HDS-drug interactions and contraindications. The findings would facilitate the health-care professionals to communicate these documented interactions and contraindications to their patients and ⁄ or caregivers thereby preventing serious adverse events and improving desired therapeutic outcomes.

Helicobacter pylori is a Gram-negative bacillus that is associated with the development of gastritis and peptic ulcer disease (PUD). In Nigeria, leaf extracts of Eucalyptus torelliana F. Muell. are used in traditional medicine to treat PUD and other gastrointestinal ailments. The additive and synergistic effects of E. torelliana leaf extracts, in combination with clarithromycin, were investigated using two types of H. pylori strains (ATCC 43629, ATCC 43579) and four clinical isolates of H. pylori (Ed, A2, G1-1, 5514) in the checkerboard assay and the fractional inhibitory concentration (FIC) index. A time-kill study was also performed on the strain ATCC 43579. The results showed that the E. torelliana extract inhibited the growth of all H. pylori strains, and the addition of one of the isolated active compounds, namely compound 2 (a substituted pyrenyl ester) enhanced the activity of clarithromycin. The minimum inhibitory concentration values of clarithromycin and the botanical compound were reduced twofold (from 0.125 to 0.0625 µg/mL and > 100 to 50 µg/mL respectively). A 100% reduction in CFU/mL of H. pylori ATCC 43579 was observed with the combination of 0.25 µg/mL clarithromycin and 100 µg/mL and 200 µg/mL compound 2 after 3 h of exposure. The results of the investigation showed that the combination of botanical compounds and antibiotics may be beneficial in the treatment of H. pylori infections.

The Dietary Supplements Information Expert Committee (DSI-EC) of the United States Pharmacopeial Convention (USP) reviews the safety of dietary supplements and dietary supplement ingredients for the purpose of determining whether they should be admitted as quality monographs into the United States Pharmacopeia and National Formulary (USP-NF). The United States Food and Drug Administration (FDA) has enforcement authority to pursue a misbranding action in those instances where a dietary supplement product indicates that it conforms to USP standards but fails to so conform. Recently DSI-EC undertook a safety evaluation of spirulina, a widely used dietary ingredient. DSI-EC reviewed information from human clinical trials, animal studies, and regulatory and pharmacopeial sources and analyzed 31 adverse event reports regarding spirulina to assess potential health concerns. At the conclusion of this review, DSI-EC assigned a Class A safety rating for Spirulina maxima and S. platensis, thereby permitting the admission of quality monographs for these dietary supplement ingredients in USP-NF. DSI-EC continually monitors reports concerning the safety of dietary supplements and dietary supplement ingredients for which USP dietary supplement monographs are developed. The DSI-EC may revisit the safety classification of spirulina as new information on this dietary ingredient becomes available.

Black cohosh [Actaea racemosa L., formerly Cimicifuga racemosa (L.) Nutt.] is an herbal medicine that has been used in the United States for over 100 years for the treatment of many conditions, including menopausal symptoms. To date the clinical data remain controversial. While there are over 20 positive clinical trials, all of the recent NIH-funded studies are negative, casting doubt on its efficacy. In addition, the cases of hepatoxicity are replete, although direct causality to the ingestion of black cohosh has not been demonstrated. In vitro studies have suggested possible chemopreventative effects; however, the in vivo studies to date do not support this hypothesis and suggest enhanced metastases. Unresolved issues of quality, safety, and efficacy remain and until these issues are resolved, the use of black cohosh products cannot be recommended.

Future research of herbal products for menopausal women should include long-term safety assessments because women may use these products for prolonged periods of time. Growing numbers of women take prescription medications and concurrently use herbal products for alleviation of menopausal symptoms. Because of possible herb-drug interactions, both drug and supplement manufacturers should provide basic pharmacokinetic data to reduce the risk of adverse interactions. In addition, herbal products produced to high quality standards are essential for ensuring consumer safety. Regulatory frameworks must be in place to ensure that herbal ingredients' identities have been verified, that they have been properly quantified per unit dose, that the product is within tolerance limits for contaminants, that the product's safety and effectiveness under the recommended conditions of use have been assessed before sale to the public, and that a system is in place to detect and deal with adverse reactions when they arise. This article explores these and related concerns.

To investigate the biological activities of Justicia pectoralis Jacq. (Acanthaceae), an herbal medicine used in Costa Rica (CR) for the management of menopausal symptoms and dysmenorrhea.The aerial parts of J. pectoralis were collected, dried and extracted in methanol. To establish possible mechanisms of action of JP for the treatment of menopausal symptoms, the estrogenic and progesterone agonists, and antiinflammatory activities were investigated.The methanol extract (JP-M) was tested in ER and PR binding assays, a COX-2 enzyme inhibition assay, the ERbeta-CALUX assay in U2-OS cells, as well as reporter and endogenous gene assays in MCF-7 K1 cells.The JP-M extract inhibited COX-2 catalytic activity (IC(50) 4.8 microg/mL); bound to both ERalpha and ERbeta (IC(50) 50 microg/mL and 23.1 microg/mL, respectively); induced estrogen-dependent transcription in the ERbeta-CALUX; and bound to the progesterone receptor (IC(50) 22.8 microg/mL). The extract also modulated the expression of endogenous estrogen responsive genes pS2, PR, and PTGES in MCF-7 cells at a concentration of 20 microg/mL. Activation of a 2 ERE-construct in transiently transfected MCF-7 cells by the extract was inhibited by the estrogen receptor antagonist ICI 182,780, indicating that the effects were mediated through the estrogen receptor. Finally, the extract weakly enhanced the proliferation of MCF-7 cells, however this was not statistically significant as compared with DMSO controls.Extracts of J. pectoralis have estrogenic, progestagenic and anti-inflammatory effects, and thus have a plausible mechanism of action, explaining its traditional use for menopause and PMS.

Black cohosh (Actaea racemosa L. [syn. Cimifuga racemosa L.]) extracts (BCE) are marketed worldwide for the management of menopausal symptoms. However, recently more than 75 cases of hepatotoxicity associated with black cohosh ingestion have been reported. While these cases have not been fully substantiated for causality, the data suggest that herb-drug interactions may be involved rather than a direct hepatotoxic event. This work describes the in vitro inhibition of four CYP450 enzymes (1A2, 2D6, 2C9, 3A4) by black cohosh extracts and identifies the active inhibitory constituents. Ethanol extracts (75 and 80% ethanol) and a 40% isopropanol extract induced a concentration-dependent inhibition of all CYP450 isozyme activities, with median inhibitory concentrations (IC(50)) ranging from 21.9 microg/ml to 65.0 microg/ml. Isolation of the active chemical constituents, showed that the triterpene glycosides were weakly active (IC(50) 25-100 microM), while fukinolic acid and cimicifugic acids A and B strongly inhibited all CYP isozymes (IC(50) 1.8-12.6 microM). None of the extracts inhibited the growth of Hep-G2 cells in concentrations up to 50 microg/ml. These data suggest that BCEs are not directly hepatotoxic, but may have the potential to induce herb-drug interactions, which may in turn explain the rare cases of hepatotoxicity observed in women using multiple medications and dietary supplements, including black cohosh.

SUMMARY Purpose The Dietary Supplements Information Expert Committee (DSI-EC; the Committee) of the United States Pharmacopeial Convention (USP) reviews safety profiles of dietary supplements before development of USP‐National Formulary (USP‐NF) quality monographs. Because the veracity of dietary supplement adverse event reports (DS AERs) directly affects DSI-EC safety reviews, the Committee reviewed the current status of DS AER reporting in the US. Methods DSI-EC reviewed PubMed searches, information from the US Food and Drug Administration’s (FDA) MedWatch program, the Toxic Exposure Surveillance System (TESS) of the American Association of Poison Control Centers (AAPCC), and reports from US and other agencies. DSI-EC analyzed this information to identify key factors that affect the quality of DS AERs. Results The overall incidence of DS AERs appears generally to be low. However, the primary reporting portal (FDA MedWatch) receives fewer AERs than do poison control centers (PCCs), and limited coordination exists among national and international surveillance programs for evaluating signals that may indicate potential public health risks. Both inadequate and poor-quality reporting of DS AERs are major limitations of DS safety monitoring in the US. Conclusions Based on its assessments, the Committee advances recommendations to improve the quality of reporting, monitoring, and assessing DS AERs. These include (1) enhanced data collection approaches, (2) improved coordination of AER surveillance programs, (3) strengthening of educational programs for public and health care sectors, and (4) conduct of research concerning the safety of DS. If taken, these approaches are expected to improve the health and well-being of DS users. Copyright # 2008 John Wiley & Sons, Ltd. key words—dietary supplements; adverse event reporting

The effect of Menoprogen (MPG) on ovarian granulosa cell (GC) apoptosis was investigated in vitro and in vivo in an aged rat model of menopause. Intragastric administration of Menoprogen or estradiol valerate to 14-month-old senile female rats for eight weeks increased plasmaE2levels, as well as the weight of both ovarian and uterine tissues. Flow cytometric (FCM) analysis of isolated GCs from MPG-treated aged rats showed reductions in theG0/G1ratio and apoptotic peaks. Isolated GCs also exhibited an increase in cell size and the number of cytoplastic organelles and intracellular gap junctions, the reappearance of secretory granules, and a lack of apoptotic bodies as determined by TEM. Results from a TdT-mediated dUTP nick end-labeling (TUNEL) assay revealed a reduction in TUNEL-positive GCs after MPG treatment. Immunohistochemical analysis showed a downregulation of proapoptotic Bax proteins and an upregulation of antiapoptotic Bcl-2 proteins. The addition of MPG-medicated serum to the media of cultured GCs also reduced cadmium chloride-induced apoptosis and downregulated caspase-3 protein expression. This work demonstrates that Menoprogen inhibits GC apoptosis in aged female rats and thereby increasesE2production. This represents a novel mechanism of action for this herbal medicine in the treatment of menopausal symptoms.

Investigation on the medical ethnobotany of the Q’eqchi Maya of Livingston, Izabal, Guatemala, was undertaken in order to explore Q’eqchi perceptions, attitudes, and treatment choices related to women’s health. Through participant observation and interviews a total of 48 medicinal plants used to treat conditions related to pregnancy, childbirth, menstruation, and menopause were collected and identified followed by the evaluation of 20 species in bioassays relevant to women’s health. Results of field interviews indicate that Q’eqchi cultural perceptions affect women’s health experiences while laboratory results (estrogen receptor and serotonin receptor binding assays) provide a scientific correlation between empirical medicinal plant use among the Q’eqchi and the pharmacological basis for their administration. These data can contribute to Guatemala’s national effort to promote a complementary relationship between traditional Maya medicine and public health services and can serve as a basis for further pharmacology and phytochemistry on Q’eqchi medicinal plants for the treatment of women’s health conditions.

The present study explored symptoms, attitudes and treatments surrounding women's health and menopause among the Q'eqchi Maya of the eastern tropical lowlands of Guatemala. Data were obtained through participant observation, semi-structured interviews, focus groups and plant walks with 50 Q'eqchi community members from the state of Izabal, Municipality of Livingston, including five midwives, five traditional male healers and eight postmenopausal women. Results indicate that the Q'eqchi Maya of Livingston possess their own cultural perceptions of women's health which affect attitudes, symptoms and treatment choices during the menopausal transition. Since discussions of menstruation and menopause are considered cultural taboos among the Q'eqchi, many women mentioned experiencing excessive preoccupation when unanticipated and unfamiliar symptoms occurred. Furthermore, many women suffered from additional hardship when their spouse misinterpreted menopausal symptoms (vaginal dryness, sexual disinterest) as infidelity. Seven of the eight postmenopausal women interviewed indicated experiencing one or more symptoms during the menopausal transition, including headaches, anxiety, muscular pain, depression, and hot flashes. These results differ from the lack of symptomatology reported in previous studies in Mexico, but are in line with the result of menopausal research conducted among other Maya groups from the highlands of Guatemala. Although the Q'eqchi did not use a specific term for "hot flash", three Q'eqchi women used the expression "baja presion" or a "lowering of blood pressure" to explain symptoms of profuse sweating followed by chills, heart palpitations, and emotional instability. The Q'eqchi Maya mentioned a number of herbal remedies to treat menopausal symptoms. Further research on these botanical treatments is needed in order to ascertain their safety and efficacy for continued use.

For many years, hormone replacement therapy (HRT) was considered the gold standard for the symptomatic treatment of menopause. Clinical trials have found that HRT reduces the symptoms of hot flashes and sweating, while also decreasing vaginal dryness and urinary tract infections. HRT has also been shown to be protective against colon cancer (37%) and hip fractures (34%). However, recent findings from the Women's Health Initiative (WHI) have revealed that long-term HRT may actually lead to an increase in heart disease (29%), breast cancer (26%) and other adverse events, such as stroke (41%). Consequently, many women in the United States and abroad are actively looking for alternative treatments for menopause, including botanical dietary supplements.

Infectious diseases are a significant cause of morbidity and mortality worldwide, accounting for approximately 50% of all deaths in tropical countries and as much as 20% of deaths in the Americas. Despite the significant progress made in microbiology and the control of microorganisms, sporadic incidents of epidemics due to drug resistant microorganisms and hitherto unknown disease-causing microbes pose an enormous threat to public health. These negative health trends call for a global initiative for the development of new strategies for the prevention and treatment of infectious disease. For over 100 years chemical compounds isolated from medicinal plants have served as the models for many clinically proven drugs, and are now being re-assessed as antimicrobial agents. The reasons for this renaissance include a reduction in the new antibacterial drugs in the pharmaceutical pipeline, an increase in antimicrobial resistance, and the need of treatments for new emerging pathogens. Literally thousands of plant species have been tested against hundreds of bacterial strains in vitro and many medicinal plants are active against a wide range of gram positive and gram negative bacteria. However, very few of these medicinal plant extracts have been tested in animal or human studies to determine safety and efficacy. This review focuses on the medicinal plants and bacteria for which there is significant published in vitro, in vivo and clinical data available. The examples provided in this review indicate that medicinal plants offer significant potential for the development of novel antibacterial therapies and adjunct treatments (i.e. MDR pump inhibitors).

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.

Due to the long-term health risks now associated with hormone replacement therapy, many menopausal women are actively seeking alternative treatments. One such alternative is black cohosh (Actaea racemosa, syn. Cimicifuga racemosa), which has been used in the United States for the treatment of gynecologic complaints for more than 100 years. Review of the published clinical data suggests that black cohosh may be useful for the treatment of menopausal symptoms, such as hot flashes, profuse sweating, insomnia, and anxiety. Results from the most recently published trial, however, indicate that black cohosh is not effective for the treatment of menopausal symptoms in breast cancer survivors being treated with tamoxifen. Because the overall quality of the published clinical trials is low, two new randomized, double-blind, placebo-controlled clinical trials are currently underway in the United States. To date, only one standardized black cohosh extract has been tested clinically; the current recommended dose is 40-80 mg per day. At least 4-12 weeks of treatment may be required before any therapeutic benefits may be apparent. Adverse reactions such as nausea, vomiting, headaches, dizziness, mastalgia, and weight gain have been observed in clinical trials. No drug interactions are reported in the medical literature. The estrogenic effects of black cohosh are controversial, and the more recent data indicate that black cohosh extracts may have an anti-estrogenic activity. Owing to potential effects on sex hormones, however, black cohosh should not be administered to children or during pregnancy and lactation.

Over the past 10 years, botanical dietary supplements containing goldenseal (Hydrastis canadensis) have become some of the best-selling products on the American market. Traditionally used by Native Americans to treat digestive complaints, goldenseal is now recommended for the treatment of common cold, influenza, menstrual disorders, and traveler's diarrhea, as well as muscular pain. In spite of these claims, there is little scientific evidence to suggest that goldenseal is effective for the prevention or treatment of any ailment. While some published data suggest that berberine, one of the active chemical constituents of goldenseal, is of some benefit for the treatment of bacterially induced diarrhea, it is not as effective as some standard antibiotics. Furthermore, considering the low concentration of berberine in goldenseal extracts and the lack of quality control over botanical dietary supplements in general, the concentration of berberine in goldenseal extracts is too low to be therapeutically effective. The safety of extended use of goldenseal extracts is also questionable, as another chemical constituent of the plant, hydrastine, may cause hypertension, exaggerated reflexes, and respiratory failure if given in sufficient doses. Furthermore, extracts of goldenseal inhibit the activity of cytochrome P450 (CYP 3A4) in vitro, and, therefore, may increase the toxicity of concomitantly administered prescription drugs t metabolized by this enzyme. Thus, due to a lack of substantiating scientific and clinical data for either safety or efficacy, the use of goldenseal is not recommended for any therapeutic indication.

Products containing Echinacea are some of the best selling and most widely used botanical dietary supplements in the United States. Commercial Echinacea extracts are manufactured primarily from 3 Echinacea species, namely Echinacea purpurea (herb, roots, or seeds), E. angustifolia (roots), and E. pallida (roots). Current recommendations for use of these products include oral administration for the prophylaxis and treatment of the common cold, bronchitis, influenza, and bacterial and viral infections of the respiratory tract. Unfortunately, much of the clinical data published prior to 1991 suffered from poor methodology and, therefore, was difficult to assess. However, more recent controlled clinical trials have suggested that while Echinacea may not be effective for the prophylaxis of upper respiratory tract infections, these products may be useful in decreasing the symptoms and duration of illness. In vitro and in vivo studies indicate that the therapeutic effects of Echinacea are due to a stimulation of cellular immune response. The major adverse events reported are allergic reactions, ranging from contact dermatitis to anaphylaxis. Patients with an allergy to plants in the daisy family (Asteraceae) should be instructed not to use products containing Echinacea. No drug interactions have been reported, however, the extracts have been shown to inhibit the activity of cytochrome P450 and, therefore, have the potential to influence drug metabolism. Echinacea-containing dietary supplements may be used safely in conjunction with antibiotics or sulfa drugs. According to the German Commission E, patients with autoimmune disorders, AIDS, HIV infection, or tuberculosis should not use Echinacea, however, there is little scientific data to substantiate these contraindications. Recommendations for oral dosage and dosage forms are difficult due to the wide variety of preparations and doses used in the clinical trials. However, based on existing data products containing pressed juice or hydroalcoholic extracts, Echinacea purpurea (leaf juice and roots) and E. pallida (roots) have the most convincing data supporting their use.

Results from randomized, controlled clinical trials support the use of standardized leaf extracts of Ginkgo biloba for the symptomatic treatment of age-associated memory impairment, dementia (including Alzheimer's disease), and intermittent claudication. Product selection is extremely important, as much of the published safety and efficacy data are based on 2 standardized extracts, both available in the US. The recommended dose of a Ginkgo biloba leaf extract (standardized to contain 24–26% flavonoid glycosides, 5–7% terpene lactones, and less than 5 parts per million ginkgolic acids) is 40 mg to 60 mg, 3 to 4 times a day. Daily administration of the extract for at least 4 to 6 weeks is required before the full therapeutic effects may be observed. While there are no known contraindications, due to potential platelet-activating factor antagonism and inhibition of platelet aggregation, Ginkgo extracts should be used cautiously in patients receiving anticoagulant or antiplatelet therapy or having bleeding disorders. A number of bleeding episodes due to co-administration of aspirin or warfarin have been reported. Other adverse reactions, such as nausea, vomiting, headaches, and dizziness have also been observed in clinical trials. Administration of Ginkgo biloba preparations to children or during pregnancy or nursing is not currently recommended.

Over the past decade, herbal medicine has become a topic of increasing global importance, with both medical and economic implications. In developing countries, as much as 80% of the indigenous populations depends on traditional systems of medicine and medicinal plants as their primary source of healthcare. Within the European Community, herbal medicines represent an important share of the pharmaceutical market, with annual sales in the range of US$7 billion. In the United States, the sale of herbal products has skyrocketed from $200 million in 1988 to >$3.3 billion in 1997. Such widespread use of botanicals throughout the world has raised serious questions concerning the quality, safety and efficacy of these products. Thus, accurate scientific assessment of herbal medicine is a prerequisite for global harmonization of herbal health claims. In 1995, as part of its overall global strategy of "Health for All" and due to numerous requests from the member states, the Traditional Medicine Program of the WHO began the extensive task of reviewing the world's scientific literature of commonly used herbal medicines and publishing this information in monographs. The WHO monographs are technical reviews of the quality, safety and efficacy of commonly used herbal medicines and are intended primarily to harmonize the proper use of herbal medicines throughout the world.

Both Korean and Siberian ginseng are used extensively for the enhancement of mental and physical performance. However, results from recent randomized, controlled clinical trials do not substantiate the purported ergogenic effects of ginseng in healthy individuals. Nevertheless, there is some evidence (both human and animal studies) to suggest that Korean and Siberian ginseng may have health promoting effects by stimulating the immune response. The recommended daily dose for Korean ginseng is 0.5 to 2 g per day of dried root or 100 to 300 mg per day of a standardized extract containing 1.5 to 7% ginsenosides and, for Siberian ginseng, 2 to 3 g per day of dried root or equivalent preparations. Patients should be cautioned not to exceed these dosages, as adverse reactions such as hypertension, diarrhea, nervousness, and insomnia are associated with excessive ingestion. For Korean ginseng, drug interactions have been reported with both phenelzine and warfarin. There is a single case report of an interaction between Siberian ginseng and digoxin; however, the clinical significance has not been established. The use of ginseng is contraindicated for children and women who are pregnant or lactating, while Siberian ginseng should also be avoided by those patients with cardiovascular disease or hypertension (BP>180/90 mmHg).

Treatment of suspension-cell cultures of Sanguinaria canadensis L. (SCP-GM) with a series of natural and synthetic retinoic acid derivatives induced benzophenanthridine alkaloid biosynthesis in a dose-dependent manner. Suspension cells were treated with one of the following: retinoic acid (RA), retinol (ROH), retinal (RHO). 13- cis-retinoic acid (cis-RA), or retinyl acetate (ROAc) in concentrations ranging from 1 to 50 μM (24 h). Each of the RA derivatives tested increased the cellular alkaloid concentrations of sanguinarine and chelerythrine by 97 to 470% (approximately 0.045–0.23% dry wt) depending on the reinoid and the dosage employed. Control SCP-GM suspension-cells accumulated only trace amounts of sanguinarine and chelerythrine, approximately 0.005% dry wt and 0.004% dry wt, respectively. The activity of S-adenosyl methionine: tetrahydroberberine-.V-methyltransferase, a key branch point enzyme in the benzophenanthridine biosynthetic pathway, was also induced over 24 h after treatment of th...

Leaf explants of Stevia rebaudiana Bertoni (Compositae), an herb which produces the sweet ent-kaurene glycoside stevioside, were cultured in Murashige and Skoog medium with vitamins, sucrose (30 g l−1), agar (0.9% w/v) and supplemented with naphthaleneacetic acid (NAA, 0.5 mg l−1) and benzylaminopurine (BAP, 0.5 mg l−1). These conditions yielded friable callus cultures. Differentiation of the callus tissue was then achieved by eliminating the agar and modulating the medium's hormone concentrations. Thus, medium containing increased auxin concentration (1.0 mg l−1) and no cytokinin or increased cytokinin (1.0 mg l−1) and no auxin yielded root or shoot cultures respectively. Supplementation of the shoot medium with NAA (1.0 mg ml−1) induced shoot cultures to grow roots thereby differentiating into rooted-shoot cultures. Only the rooted-shoot cultures tasted sweet. Feedings of [2-14C]acetic acid to callus, shoot or rooted-shoot cultures demonstrated that only the rooted-shoot cultures are capable of de novo biosynthesis of the aglycone moiety of stevioside (steviol). In addition, [methyl-3H(N)steviol feedings to shoot or rooted-shoot cultures illustrated that both types of cultures are capable of the glycosylation reaction. The ability of these tissues to glycosylate steviol to stevioside was also demonstrated employing crude enzyme preparations derived from shoot or rooted-shoot cultures. These results suggest that stevioside biosynthesis is a function of tissue differentiation since both roots and leaves are required for cultured S. rebaudiana to biosynthesize stevioside from acetate, while the final biosynthetic steps can be performed at all levels of differentiation.

Since the publication of the results of the Women’s Health Initiative that described the risks of hormone replacement therapy, many women are actively seeking alternative treatments for menopausal symptoms. Black cohosh (Actaea racemosa, syn. Cimicifuga racemosa) is one such alternative that has been used in the US for over 100 years. To date only two cimicifuga extracts have been tested clinically, and the current recommended dosage is 40–80 mg/day. Review of the published clinical data suggests that cimicifuga may be useful for the treatment of menopausal symptoms, such as hot flashes, profuse sweating, insomnia, and anxiety. However, the methodology used in most of the trials is poor and further clinical assessment of cimicifuga is needed. In terms of safety, transient adverse events such as nausea, vomiting, headaches, dizziness, mastalgia, and weight gain have been observed in clinical trials. A few cases of hepatotoxicity have been reported, but a direct association with the ingestion of cimicifuga has not been demonstrated. The most recent data suggest that cimicifuga is not estrogenic.

Many women seek safe and effective alternative therapies for the treatment of menopausal symptoms. Black cohosh, a botanical product, is one such treatment. Its mechanism of action may involve estrogenic effects, but new data dispute the estrogenic theory and indicate that extracts of black cohosh do not bind to the estrogen receptor, up-regulate estrogen-dependent genes, or stimulate the growth of estrogen-dependent tumors in animal models.

A simple and relatively rapid technique is described which can be used to determine accurately the overall pattern of macromolecule biosynthesis in plant cells. Sanguinaria canadensis L. (Papaveraceae) suspension cells were used as the model system. The patterns of DNA, RNA and protein synthesis were determined through cellular incorporation of [methyl-3H]-thymidine, [5-3H]-uridine and [4,5-3H]-leucine, respectively. The method takes advantage of a one-step filtering procedure, which simplifies the extraction methods normally used. The results are described and possible applications of this procedure in plant-cell growth studies and biotechnology are discussed.