Natural Estrogenic Compounds Research Articles

G Protein-Coupled Estrogen Receptor GPER: Molecular Pharmacology and Therapeutic Applications.

The actions of estrogens and related estrogenic molecules are complex and multifaceted in both sexes. A wide array of natural, synthetic, and therapeutic molecules target pathways that produce and respond to estrogens. Multiple receptors promulgate these responses, including the classical estrogen receptors of the nuclear hormone receptor family (estrogen receptors α and β), which function largely as ligand-activated transcription factors, and the 7-transmembrane G protein-coupled estrogen receptor, GPER, which activates a diverse array of signaling pathways. The pharmacology and functional roles of GPER in physiology and disease reveal important roles in responses to both natural and synthetic estrogenic compounds in numerous physiological systems. These functions have implications in the treatment of myriad disease states, including cancer, cardiovascular diseases, and metabolic disorders. This review focuses on the complex pharmacology of GPER and summarizes major physiological functions of GPER and the therapeutic implications and ongoing applications of GPER-targeted compounds.

Natural and synthetic estrogenic compounds in the Pearl River Estuary and northern shelf of the South China Sea

Endocrine disrupting compounds and in particular estrogenic substances have the ability to interact with the hormone system of organisms. Among them are not only synthetic but also natural substances that potentially stress the aquatic ecosystem. High human population densities such as around the Pearl River Estuary (PRE) are suspected of exerting significant anthropogenic pressure onto coastal areas. In this study, natural and synthetic estrogens as well as estrogen-like substances derived from plants and fungi were investigated in the PRE and at the adjacent northern shelf of the South China Sea. Maximum concentration of 3.6 ng L−1 for estrone (E1), 0.7 ng L−1 for 17α-ethinylestradiol (EE2), 12.9 ng L−1 for genistein (GEN), 11.9 ng L−1 for daidzein (DAI) and 1.9 ng L−1 for zearalenone (ZEN) were observed. While E1 and EE2 were detected in fresh and saltwater samples, GEN, DAI and ZEN were observed only at freshwater sampling sites. During the investigations, the analysis of 17β-estradiol (E2) and EE2 indicated a strong matrix dependence. Additionally, an estrogen screen observation showed estrogenic activity in form of estradiol equivalent quotients up to 0.18 ng L−1.

Assessment of environmental applicability of TiO2 coated self-cleaning glass for photocatalytic degradation of estrone, 17β-estradiol and their byproducts

Optimization of photocatalytic degradation of two natural estrogenic compounds, estrone (E1) and 17β-estradiol (17β-E2) in aqueous medium was performed on TiO2 coated Pilkington ActivTM self-cleaning glass as a novel approach to eliminate free nano-TiO2 releasing to the intended environment after treatment. The active glass was characterized by Atomic Force Microscopy (AFM), X-ray diffraction (XRD), and Raman spectroscopy to characterize the TiO2 nanoparticles. The main purposes were mineralization of target compounds in the treated water during the photocatalytic reaction and also to investigate the oxidation by products. Response Surface Methodology (RSM) has been applied to optimize the photocatalytic degradation by changing time, pH, and light intensity as effective factors. According to the results, time was the more effective parameter. The maximum efficiency degradation was achieved in alkaline media. Due to interactive effects between variable factors, 1 mg/L aqueous solution of E1 and 17β-E2 in water was totally decomposed by TiO2 photocatalyzed reactions under UV-C irradiation of 10.08 W/m2 for 52.49 min at pH 9.42. Results of GC-MS analysis were introduced 17-deoxy Estrone and 2-Hydroxyestradiol as intermediate products for E1 and 17β-E2, respectively. All of the peaks finally disappeared after 170 min. Optimized conditions were applied for real sample of wastewater, presenting 30.40% and 56.84% in the efficiency degradation of E1 and 17β-E2, respectively.

An object-oriented Bayesian network approach for establishing swine manure-borne natural estrogenic compounds budget

A facility-wide estrogen budget model was developed to assess the excretion of natural estrogens by swine in a commercial swine farrowing concentrated animal feeding operations (CAFO) in North Carolina, using an object-oriented Bayesian network (OOBN) approach. The OOBN model is the combination of twelve objects of Bayesian network models, which characterize the estrogen budget flows based on the sow reproductive cycle (i.e., pre-estrus, estrus, and lactation) for the three natural estrogen types [estrone (E1), 17β-estradiol (E2), and estriol (E3)] within each barn. This OOBN model provides a mechanism to quantify the levels of the natural estrogens and their probabilistic distributions with regard to estrogen type, waste sources such as urine, feces, and recycling lagoon slurry, and animal reproductive status. Moreover, the OOBN model allows us to assess the overall contribution of natural estrogen compounds from each operational unit of the CAFO, while accounting for the uncertainties. Results from the OOBN model indicate a rank order of lactating sows > gestating sows > breeding sows in terms of contribution of estrogen loads to the total natural estrogen budget. As to estrogen type, E1 was found as the major estrogen metabolite with the summed concentrations of urine, feces, and flushing slurry wastes exceeding 3000 ng/l > 90% of the time. As to waste sources, the flushing slurry waste was found to be a major contributor of the estrogen budget compared with urine and feces wastes from barn animals.

Fitoöstrojenler ve Üreme Sağlığı

Fitoöstrojenler doğal veya yapay östrojenik bileşiklere benzer kimyasal yapılara sahip bitkiler tarafından üretilen steroid olmayan bileşiklerdir. Fitoöstrojenler, östrojene spesifik reseptörlere affinite göstererek zayıf östrojenik etki göstermektedirler. Bu nedenle üreme sağlığı üzerine etkileri konusundaki çalışmalar gün geçtikçe yoğunlaşmaktadır. Bu derleme makale, pek çok fizyolojik rolü olduğu bilinen fitoöstrojenlerin kadın ve erkek üreme sağlığı üzerindeki etkilerini belirlemek amacıyla hazırlanmıştır. Çalışmalar, özellikle yenidoğan dönemde fitoöstrojenlere (soya-izoflavon) dikkatle yaklaşılması gerektiğini, ileriki dönemlerde özellikle kızlarda seksüel dimorfik davranışları ve gelişimsel süreçlerini etkileyebileceğini göstermektedir. Aynı zamanda, fitoöstrojenlerin erken puberte başlangıcı ile ilişkili olabileceği, premenopozal dönemde mensturasyon siklusunda düzensizliklere neden olabileceği, postmenopozal dönemde ise semptomların azalmasını sağlayarak olumlu etkileri bulunabileceği görülmüştür. Erkeklerde ise fitoöstrojenlerin üreme sağlığı üzerine etkisi bulunmamıştır. Memelilerin gelişimsel süreçleri göz önünde bulundurulduğunda, evrelere göre fitoöstrojenlerin üreme sağlığı üzerine olumlu ya da olumsuz sayılabilecek önemli etkileri bulunmaktadır. Fitoöstrojenlerin etkileri üzerine insan çalışmaları sınırlı olsa da çalışmaların sonuçları yapılan pek çok sayıdaki hayvan çalışmaları ile benzerlik göstermektedir. Ancak, çalışmalardaki yöntemsel farklılıklar sonucu ortaya çıkan çelişkili sonuçlar fitoöstrojenlerin üreme sağlığı üzerindeki etkisini değerlendirmeyi güçleştirmektedir.

Estrogens and Androgens in Skeletal Physiology and Pathophysiology.

Estrogens and androgens influence the growth and maintenance of the mammalian skeleton and are responsible for its sexual dimorphism. Estrogen deficiency at menopause or loss of both estrogens and androgens in elderly men contribute to the development of osteoporosis, one of the most common and impactful metabolic diseases of old age. In the last 20 years, basic and clinical research advances, genetic insights from humans and rodents, and newer imaging technologies have changed considerably the landscape of our understanding of bone biology as well as the relationship between sex steroids and the physiology and pathophysiology of bone metabolism. Together with the appreciation of the side effects of estrogen-related therapies on breast cancer and cardiovascular diseases, these advances have also drastically altered the treatment of osteoporosis. In this article, we provide a comprehensive review of the molecular and cellular mechanisms of action of estrogens and androgens on bone, their influences on skeletal homeostasis during growth and adulthood, the pathogenetic mechanisms of the adverse effects of their deficiency on the female and male skeleton, as well as the role of natural and synthetic estrogenic or androgenic compounds in the pharmacotherapy of osteoporosis. We highlight latest advances on the crosstalk between hormonal and mechanical signals, the relevance of the antioxidant properties of estrogens and androgens, the difference of their cellular targets in different bone envelopes, the role of estrogen deficiency in male osteoporosis, and the contribution of estrogen or androgen deficiency to the monomorphic effects of aging on skeletal involution.

Anthropogenic pressure in a Portuguese river: Endocrine-disrupting compounds, trace elements and nutrients

ABSTRACTNatural organic compounds such as phytoestrogens and phytosterols found in various plants, as well as mycotoxins produced by fungi, can be found in aquatic environments. The aim of this study was to investigate the occurrence of three different classes of natural estrogenic compounds, i.e., phytoestrogens, phytosterols and mycotoxins, in estuarine water samples from the Ave River estuary. For that, water samples were collected at five sampling points distributed along the estuary at low tide, during 1 year, processed by solid-phase extraction (SPE) and analyzed by gas chromatography coupled to mass spectrometry (GC–MS). To correlate the presence of phytoestrogens and phytosterols in the estuarine water, local flora was collected on riverside. Trace elements content and physicochemical parameters such as nutrients and dissolved oxygen were also determined seasonally at each sampling point, to give insights for the evaluation of water quality and anthropogenic pressure. Both phytoestrogens and phytosterols showed a seasonal variation, with the highest values observed in spring and summer and the lowest in winter. Daidzein (DAID) was found up to 404.0 ng L−1 in spring and coumestrol (COUM) was found up to 165.0 ng L−1 in summer. The mycotoxin deoxynivalenol (DON) was ubiquitously determined with values ranging from 59.5 to 642.4 ng L−1. Nutrients and metals distribution and concentration varied among sampling stations and seasons. This study revealed for the first time the presence of mycotoxins, various classes of phytoestrogens and stigmasterol (STG) in estuarine water from the Ave River (Portugal), and the evaluation of the water quality confirmed that this estuary is still highly impacted by anthropogenic activities.

Involvement of Estrogen Receptors in the Anxiolytic-Like Effect of Phytoestrogen Genistein in Rats with 12-Weeks Postovariectomy

Phytoestrogens are natural compounds found in some vegetables, and they replicate many of the physiochemical and physiological properties of estrogens, including the regulation of mood. The phytoestrogen genistein exerts anxiolytic-like effects in rats with a chronic absence of ovarian hormones, but the mechanism involved in this effect remains to be explored. The present study explored the participation of estrogen receptor-β in the anxiolytic-like effect of genistein (1.0 mg/kg, i.p., for 4 days) in Wistar rats with 12-weeks postovariectomy, considered as experimental model of post-surgical menopause. In the light/dark test, a useful tool for anxiety study and for the screening of anxiolytic drugs, genistein reduced the latency to enter and increased the time spent in the light compartment and significantly increased the frequency and time spent exploring the light compartment compared with the control group, which is considered as an anxiolytic-like effect at experimental level. All behavioral effects produced by genistein in the light/dark test were blocked by previous tamoxifen administration (5.0 mg/kg, s.c., for 6 days), a non selective antagonist for estrogen receptor-β. The effects produced by genistein or tamoxifen in this test were not related to significant changes in general motor activity evaluated in the open field test. In conclusion, the specific contribution of present investigation was identify that estrogen receptor-β is involved in the anxiolytic-like effect produced by phytoestrogen genistein in rats with a long-term absence of ovarian hormones; supporting the hypothesis that estrogen receptor-β participates in the regulation of anxiety associated with low concentration of ovarian hormones and in the anxiolytic-like effects produced by natural estrogenic compounds such as phytoestrogens.

Identification and characterization of a novel estrogenic ligand actinopolymorphol A

Xenoestrogenic compounds are abundant in the modern environment including phytoestrogens from plants, chemical by-products from industry, and secondary metabolites from microbes; all can profoundly affect human health. Consequently mechanism-based screens are urgently needed to improve the rate at which the xenoestrogens are discovered. Estrogen Receptor (ER) dimerization is required for target gene transcription. The three ER dimer pairs (ERα/α homodimers, ERβ/β homodimers, and ERα/β heterodimers) exhibit diverse physiological responses in response to ligand-dependent activation with ERα/α homodimers being pro-proliferative and ERβ/β homodimers being anti-proliferative. The biological role of the ERα/β heterodimer remains unclear. We previously developed a cell-based, bioluminescence resonance energy transfer (BRET) assay that can distinguish natural estrogenic compounds based on their abilities to activate the three diverse ER dimer pairs. Using BRET assays, we sought to identify novel xenoestrogens from soil bacteria that preferentially activate ERα/β heterodimer with hopes of shedding light on the biological function of this elusive dimer pair. Here we describe the application of BRET assays in high throughput screens of crude bacterial extracts not previously screened for ER modulatory function and originating from unique ecological niches. Here we report the discovery and biological evaluation of a new natural product, actinopolymorphol A (1), that preferentially induces ERα/β dimerization. Actinopolymorphol A represents the first representative of a new ER modulatory scaffold.

Determination of estrogenic compounds in wastewater using liquid chromatography–tandem mass spectrometry with electrospray and atmospheric pressure photoionization following desalting extraction

Two complementary LC–MS ionization methods, electrospray (ESI) and atmospheric pressure photoionization (APPI), have been optimized to determine three natural estrogenic compounds (estrone, 17β-estradiol and estriol) and two synthetic estrogenic compounds (17α-ethynylestradiol and diethylstilbestrol) in the influent and effluent of wastewater treatment plants (WWTPs). The wastewater samples were first subjected to solid-phase extraction coupled with desalting extraction to remove matrix interference. The analytes were then detected using liquid chromatography–tandem mass spectrometry (LC–MS–MS) with ESI and dopant-assisted (DA) APPI to evaluate the ion suppression effect and to complement the detection and quantification of estrogenic compounds in complex wastewater samples. The average ion suppression factors for the extracts of the WWTP influent analyzed using ESI and APPI were 52 ± 5% and 27 ± 7%, respectively. The sensitivity and ionization efficiency of the LC–ESI–MS–MS system decreased dramatically when a complex matrix was present in the WWTP influent sample. Estrogenic compounds could be detected in the WWTP influent and effluent samples at concentrations below the parts-per-billion level. The lower detection limits obtained when using ESI and the higher matrix tolerance of the APPI method allowed the complete quantification of estrogenic compounds in very complex samples in a complementary manner.

Modeling coupled degradation, sorption, and transport of 17β‐estradiol in undisturbed soil

The presence of 17β‐estradiol in the environment, even at parts‐per‐trillion concentrations, raises significant concern regarding the health of aquatic organisms. Once 17β‐estradiol is released into the environment from human and animal sources, its fate and transport is controlled by factors such as sorption and transformation, which need to be understood to fully assess potential exposures. The objective of this study was first to discern, through controlled batch experiments, the simultaneous transformation (i.e., chemical and biological) of natural estrogenic compounds and their mass exchange between the aqueous and solid phase (i.e., reversible and irreversible sorption sites). In addition, a model was developed that used a series of first‐order expressions to describe the various fate and transport processes of parent and metabolite estrogens in the nonequilibrium batch experiments. A global optimization method was used to estimate the parameters of this nonequilibrium batch model. The model provided a good description of the data, and the parameter estimates were reliable. The batch studies parameter estimates were then incorporated into a convective‐dispersive model to describe two undisturbed column experiments. The consistency of parameter estimates between the batch and column experiments indicated a high capability and reliability of this model and the parameter values.

Removal of Selected Natural and Synthetic Estrogenic Compounds in a Canadian Full‐Scale Municipal Wastewater Treatment Plant

The effect of a full-scale municipal wastewater treatment plant (WWTP) and each of the treatment units within the stream on the removal of endocrine-disrupting compounds was evaluated by tracking 17-beta-estradiol (E2), estrone (E1), and 17-alpha-ethinylestradiol (EE2). The overall performance of the WWTP compared well with other plants, as 90.5% removal of E1+E2 and 74.9% removal of EE2 were observed. A larger fraction of EE2 entered the plant in particulate form than E1 and E2, while a lower fraction of EE2 left the plant in particulate form than soluble form. The activated sludge units reduced the concentration of E1+E2 and EE2 in the liquid phase by 88.2% and 44.6%, respectively. The UV treatment process did not reduce the amount of estrogens. The aqueous phase of the tertiary lagoon solids contained higher levels of estrogens compared with the lagoon influent.

Functional SNPs of the breast cancer resistance protein ‐ therapeutic effects and inhibitor development

Breast cancer resistance protein (BCRP) is a half-molecule ATP-binding cassette transporter that pumps out various anticancer agents such as 7-ethyl-10-hydroxycamptothecin, topotecan and mitoxantrone. We have previously identified three polymorphisms within the BCRP gene, G34A (substituting Met for Val-12), C376T (substituting a stop codon for Gln-126) and C421A (substituting Lys for Gln-141). C421A BCRP-transfected murine fibroblast PA317 cells showed markedly decreased protein expression and low-level drug resistance when compared with wild-type BCRP-transfected cells. In contrast, G34A BCRP-transfected PA317 cells showed a similar protein expression and drug resistance profile to wild-type. The C376T polymorphism would be expected to have a considerable impact as active BCRP protein will not be expressed from a T376 allele. Hence, people with C376T and/or C421A polymorphisms may express low levels of BCRP, resulting in hypersensitivity of normal cells to BCRP-substrate anticancer agents. Estrogens, estrone and 17β-estradiol, were previously found to restore drug sensitivity levels in BCRP-transduced cells by increasing the cellular accumulation of anticancer agents. BCRP transports sulfated estrogens but not free estrogens and in a series of screening experiments for synthesized and natural estrogenic compounds, several tamoxifen derivatives and phytoestrogens/flavonoids were identified that effectively circumvent BCRP-mediated drug resistance. The kinase inhibitors gefitinib and imatinib mesylate also interact with BCRP. Gefitinib, an inhibitor of epidermal growth factor receptor-tyrosine kinase, inhibits its transporter function and reverses BCRP-mediated drug resistance both in vitro and in vivo. BCRP-transfected human epidermoid carcinoma A431 cells and BCRP-transfected human non-small cell lung cancer PC-9 cells show gefitinib resistance. Imatinib, an inhibitor of BCR-ABL tyrosine kinase, also inhibits BCRP-mediated drug transport. Hence, both functional SNPs and inhibitors of BCRP reduce its transporter function and thus modulate substrate pharmacokinetics and pharmacodynamics.

Expression of the estrogen-inducible EGFP gene in aromatase-null mice reveals differential tissue responses to estrogenic compounds

Aromatase is an enzyme responsible for the conversion of androgen to estrogen. We genetically engineered an aromatase-deficient mouse (Ar −/− mouse) to express an enhanced green fluorescent protein (EGFP) gene in the uterus, ovary, adrenal and pituitary glands in a 17β-estradiol (E2)-inducible manner. In this study, we analyzed estrogenic activities of diethylstilbestrol, genistein, daidzein and E2 in the Ar −/− tissues by using the EGFP expression as an indicator. These analyses manifest differential responses of the tissues to the compounds and also allow to determine the relative estrogenic potency of the compounds to that of E2 in vivo. Furthermore, analyses of the EGFP expression in ERα-deficient mice suggested that the expression is ERα-dependent in the uterus and pituitary gland. In conclusion, the Ar −/− mouse carrying the E2-inducible EGFP gene is a valuable tool for quantitative analyses of natural and synthetic estrogenic compounds in vivo.

Determination of natural and synthetic estrogenic compounds in coastal lagoon waters by HPLC-electrospray-mass spectrometry

A new analytical method was developed for the simultaneous determination of estrogenic compounds of natural (estradiol, estriol, estrone) and synthetic origin, both steroidal (ethinylestradiol, mestranol) and non-steroidal (benzophenone, bisphenol-A, diethylstilbestrol, octylphenol, nonylphenol, nonylphenol monoethoxylate carboxylate), in environmental aqueous samples by high-performance liquid chromatography coupled with ion trap-mass spectrometry via electrospray interface (HPLC-ESI-IT-MS). Quantitative MS detection was performed in the negative mode for all compounds except mestranol and benzophenone, which were detected under positive ion conditions. Very low method detection limits (MDLs), between 0.1 and 2.6 ng/L, were achieved in coastal lagoon water samples, while the developed solid-phase-extraction (SPE) procedure permitted simultaneous recovery of all analytes from spiked water samples with yields >70% (7–11 RSD%), except estriol and benzophenone, which were recovered with 60% (9 RSD%) and 50% (11 RSD%) yields, respectively. The proposed method was applied to the analysis of Venice (Italy) lagoon waters, where average concentrations of selected compounds in the 2.8–33 ng/L concentration range were found.

Estrogenic potential of the Venice, Italy, lagoon waters.

The exposure of the Venice lagoon (Italy) to endocrine-disrupting compounds (EDCs) from different sources was investigated. Spatial and time distribution of EDC concentrations were determined in four sampling sessions (December 2001-May 2002) by solid phase extraction followed by high-performance liquid chromatography separation coupled with mass spectrometry detection via electrospray interface (SPE-HPLC-ESI-MS), which allowed identification of natural (estradiol, estrone) and synthetic estrogenic compounds, both steroidal (ethinylestradiol, mestranol) and nonsteroidal (benzophenone, bisphenol-A, nonylphenol, nonylphenol monoethoxylate carboxylate). No significant differences in the EDC distribution were observed between stations located near selected sources (raw sewage from the historical center of Venice, treated municipal and industrial effluents from sewage treatment plants, and areas undergoing the inflow of rivers). While synthetic nonsteroidal analytes were recorded in the 1 to 1040 ng/L range (average concentration: 34 ng/L), steroidal EDC (estradiol, ethinylestradiol) concentrations were lower (1-125 ng/L; average concentration: 8 ng/L). The estrogenic activity of lagoon waters was estimated in terms of estradiol equivalent concentration (EEQ) by applying the estradiol equivalency factors (EEFs). Steroidal EDCs (estradiol, ethinylestradiol) contributed >97% to the total potential estrogenicity of the waters, which accounted for 4 to 172 ng/L (average: 25 ng/L), as total EEQs. These levels are likely to pose adverse effects on the Venice lagoon aquatic organisms.

Identification of Estrogen-Responsive Genes by Complementary Deoxyribonucleic Acid Microarray and Characterization of a Novel Early Estrogen-Induced Gene:EEIG1

Estrogen receptors (ERs) are nuclear transcription factors that regulate gene expression in response to estrogen and estrogen-like compounds. Identification of estrogen-regulated genes in target cells is an essential step toward understanding the molecular mechanisms of estrogen action. Using cDNA microarray examinations, 19 genes were identified as induced by 17 beta-estradiol in MCF-7 cells, 10 of which have been reported previously to be estrogen responsive or to be linked with ER status. Five known estrogen-regulated genes, E2IG4, IGFBP4, SLC2A1, XBP1 and B4GALT1, and AFG3L1, responded quickly to estrogen treatment. A novel estrogen-responsive gene was identified and named EEIG1for early estrogen-induced gene 1. EEIG1 was clearly induced by 17 beta-estradiol within 2 h of treatment, and was widely responsive to a group of estrogenic compounds including natural and synthetic estrogens and estrogenic environmental compounds. EEIG1 was expressed in ER-positive but not in ER-negative breast cancer cell lines. EEIG1 expression was repressed by antiestrogens 4-OH-tamoxifen and ICI 182,780 but not by protein synthesis inhibitors cycloheximide and puromycin. These results provide evidence that some estrogenic compounds differentially enhance the transcription of estrogen-regulated genes and suggest a role for EEIG1 in estrogen action.

Differential activation of wild-type and variant forms of estrogen receptor α by synthetic and natural estrogenic compounds using a promoter containing three estrogen-responsive elements

Structure-dependent estrogen receptor α (ERα) agonist and antagonist activities of synthetic and natural estrogenic compounds were investigated in human HepG2, MDA-MB-231 and U2 cancer cell lines. Compounds used in this study include 4′-hydroxytamoxifen, ICI 182,780, bisphenol-A (BPA), 2′,4′,6′-trichloro-4-biphenylol (3Cl-PCB-OH), 2′,3′,4′,5′-tetrachloro-4-biphenylol (4Cl-PCB-OH), p-t-octylphenol, p-nonylphenol, naringenin, kepone, resveratrol, and 2,2-bis( p-hydroxyphenyl)-1,1,1-trichloroethane (HPTE). Cells were transfected with a construct (pERE 3) containing three tandem estrogen responsive elements (EREs) and either wild-type estrogen receptor α (ER-wt) or variants expressing activation function-1 (ER-AF1) or AF-2 (ER-AF2). The ER agonist activities of the synthetic mono and dihydroxy aromatic compounds are comparable in all three-cell lines, whereas the activities of naringenin, kepone and resveratrol are dependent on cell context and expression of wild-type or variant forms of ERα. In contrast, the ER antagonist activities for these compounds were highly complex and, with the exception of 3Cl-PCB-OH, all compounds inhibited E2-induced wild-type or variant ER action. Results of this in vitro study suggest that the estrogenic and antiestrogenic activity of structurally diverse synthetic and natural estrogenic compounds is complex, and this is consistent with published data that often give contradictory results for these compounds.

Medicinal foodstuffs. XVIII. Phytoestrogens from the aerial part of Petroselinum crispum MIll. (Parsley) and structures of 6"-acetylapiin and a new monoterpene glycoside, petroside.

In the course of our screening for natural estrogenic compounds from Occidental medicinal herbs, the extracts of several herbs were found to show proliferative activity in MCF-7 (an estrogen-sensitive breast cancer cell line). Among these active herbs, the methanolic extract from the aerial parts of Petroselinum crispum (parsley) showed potent estrogenic activity, which was equal to that of isoflavone glycosides from soybean. Through bioassay-guided separation, we isolated several flavone glycosides and a new flavone glycoside, 6"-acetylapiin, with estrogenic activity together with a new monoterpene glucoside, petroside. The structures of 6"-acetylapiin and petroside were characterized by the chemical and physicochemical evidence. Estrogenic activities of these flavone glycosides were found to be enhanced by removal of their glycoside moieties. The EC50 values (concentration needed to enhance the MCF-7 proliferation 50% compared to non-estrogen treated cell) of their aglycones are as follows, apigenin (1.0 microM), diosmetin (2.9 microM), and kaempferol (0.56 microM). The estrogenic activities of these flavones are nearly equal to those of the isoflavones, daidzein (0.61 microM) and genistein (0.60 microM). The methanolic extract of parsley, apiin, and apigenin restored the uterus weight in ovariectomized mice when orally administered for consecutive 7 days.

Methods for xenoestrogen testing

Research in our laboratories has focused on development of a battery of in vivo and in vitro bioassays for determining estrogenic activity and potency of different classes of natural and synthetic industrial-derived estrogenic compounds (xenoestrogens) including food/beverage extracts, phytoestrogens, phenolic compounds, organochlorine pesticides and pollutants. For many of the weak estrogenic compounds, their activity as estrogen receptor (ER) agonists or antagonists is dependent on the gene/gene promoter, cell context and expression of ERα or ERβ isoform. For example, extracts of red wine, bound to the ER, exhibited estrogenic activity in T47D, MCF-7 (breast) and Hep G2 (liver) human cancer cell lines, whereas reconstituted organochlorine pesticide residues found in food were active only in Hep G2 cells that transiently expressed ERα or ERβ. The relative potencies of red wine extracts versus reconstituted organochlorine pesticides were assay-dependent; however, estrogen equivalent daily intakes from a glass of red wine (∼0.5–2 μg estrogen equivalents/day) were at least 103 higher than observed for the reconstituted organochlorine pesticide mixtures. Risk assessment of xenoestrogens and other synthetic chemicals which modulate endocrine responses must take into account high dietary levels of natural products in food, drugs and health food store extracts which also modulate endocrine responses.