Novel BODIPY-estradiol conjugates have been synthesized by selecting position C-3-O for labeling. The conjugation strategy was based on Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC) or etherification. Estradiol derivatives used as azide partners bearing an ω-azidoalkyl function through C4-C8-long linkers have been prepared. CuAAC reactions of estradiol azides with BODIPY alkyne furnished fluorescent 3-O-labeled conjugates bearing the triazole ring as a coupling moiety. Williamson etherifications of 3-O-(ω-bromoalkyl)-17β-estradiol derivatives with BODIPY-OH resulted in labeled conjugates connected with an ether moiety. Interactions of the conjugates with estrogen receptor (ER) were investigated using molecular docking calculations in comparison with estradiol. The conjugates occupied both the classical and alternative binding sites on human ERα, with slightly lower binding affinity to references estradiol and diethystilbestrol. All compounds have displayed reasonable estrogenic activity. They increased the proliferation of ER-positive breast cancer cell line MCF7 contrary to ER-negative SKBR-3 cell line. The most potent compound 13a induced the transcriptional activity of ER in dose-dependent manner in dual luciferase recombinant reporter model and increased progesterone receptor's expression, proving the retained estrogenic activity. The fluorescence of candidate compound 13a co-localised with the ERα. The newly synthesized labeled compounds might serve as good starting point for further development of fluorescent probes for modern biological applications. In addition to studying steroid uptake and transport in cells, e.g. in the processes of biodegradation of estrogen-hormones micropollutants, they could also be utilized in examination of estrogen-binding proteins.

• Klíčová slova
• 17β-estradiol, BODIPY, CuAAC, Estrogenic activity, Human estrogen receptor alpha,
• MeSH
• alfa receptor estrogenů * metabolismus   chemie   MeSH
• azidy chemie   MeSH
• estradiol * chemie   farmakologie   MeSH
• estrogeny chemie   MeSH
• fluorescenční barviva chemie   MeSH
• lidé MeSH
• MFC-7 buňky MeSH
• nádorové buněčné linie MeSH
• proliferace buněk účinky léků   MeSH
• simulace molekulového dockingu * MeSH
• sloučeniny boru * chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene MeSH Prohlížeč
• alfa receptor estrogenů * MeSH
• azidy MeSH
• ESR1 protein, human MeSH Prohlížeč
• estradiol * MeSH
• estrogeny MeSH
• fluorescenční barviva MeSH
• sloučeniny boru * MeSH

Mixture toxicity, including agonistic and antagonistic effects, is an unrevealed environmental problem. Estrogenic endocrine disruptors are known to cause adverse effects for aquatic biota, but causative chemicals and their contributions to the total activity in sewage sludge remain unknown. Therefore, advanced analytical methods, a yeast bioassay and mixture toxicity models were concurrently applied for the characterization of 8 selected sludges with delectable estrogenic activity (and 3 sludges with no activity as blanks) out of 25 samples from wastewater treatment plants (WWTPs). The first applied full logistic model adequately explained total activity by considering the concentrations of the monitored compounds. The results showed that the activity was primarily caused by natural estrogens in municipal WWTP sludge. Nevertheless, activity in a sample originating from a car-wash facility was dominantly caused by partial agonists - nonylphenols - and only a model enabling prediction of all dose-response curve parameters of the final mixture curve explained these results. Antiestrogenic effects were negligible, and effect-directed analysis identified the causative chemicals.

• Klíčová slova
• Agonistic activity, Antagonistic activity, Constructed wetland, Mixture toxicity, WWTP sludge,
• MeSH
• antagonisté estrogenu MeSH
• chemické látky znečišťující vodu * analýza   toxicita   MeSH
• endokrinní disruptory * analýza   toxicita   MeSH
• estrogeny toxicita   MeSH
• odpadní vody MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antagonisté estrogenu MeSH
• chemické látky znečišťující vodu * MeSH
• endokrinní disruptory * MeSH
• estrogeny MeSH
• odpadní vody MeSH

Stagnant freshwaters can be affected by anthropogenic pollution and eutrophication that leads to massive growth of cyanobacteria and microalgae forming complex water blooms. These can produce various types of bioactive compounds, some of which may cause embryotoxicity, teratogenicity, endocrine disruption and impair animal or human health. This study focused on potential co-occurrence of estrogenic and retinoid-like activities in diverse stagnant freshwaters affected by phytoplankton blooms with varying taxonomic composition. Samples of phytoplankton bloom biomass and its surrounding water were collected from 17 independent stagnant water bodies in the Czech Republic and Hungary. Total estrogenic equivalents (EEQ) of the most potent samples reached up to 4.9 ng·g-1 dry mass (dm) of biomass extract and 2.99 ng·L-1 in surrounding water. Retinoic acid equivalent (REQ) measured by in vitro assay reached up to 3043 ng·g-1 dm in phytoplankton biomass and 1202 ng·L-1in surrounding water. Retinoid-like and estrogenic activities at some sites exceeded their PNEC and effect-based trigger values, respectively. The observed effects were not associated with any particular species of cyanobacteria or algae dominating the water blooms nor related to phytoplankton density. We found that taxonomically diverse phytoplankton communities can produce and release retinoid-like compounds to surrounding water, while estrogenic potency is likely related to estrogens of anthropogenic origin adsorbed to phytoplankton biomass. Retinoids occurring in water blooms are ubiquitous signalling molecules, which can affect development and neurogenesis. Selected water bloom samples (both water and biomass extracts) with retinoid-like activity caused effects on neurodifferentiation in vitro corresponding to those of equivalent all-trans-retinoic acid concentrations. Co-occurrence of estrogenic and retinoid-like activities in stagnant water bodies as well as the potential of compounds produced by water blooms to interfere with neural differentiation should be considered in the assessment of risks associated with water blooms, which can comprise complex mixtures of natural and anthropogenic bioactive compounds.

• Klíčová slova
• Endocrine disruption, Estrogenic activity, Neural differentiation, Phytoplankton water bloom, Retinoid-like activity,
• MeSH
• estrogeny analýza   MeSH
• estron MeSH
• eutrofizace MeSH
• fytoplankton MeSH
• komplexní směsi MeSH
• lidé MeSH
• retinoidy * MeSH
• rostlinné extrakty MeSH
• sinice * MeSH
• tretinoin MeSH
• voda MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• estrogeny MeSH
• estron MeSH
• komplexní směsi MeSH
• retinoidy * MeSH
• rostlinné extrakty MeSH
• tretinoin MeSH
• voda MeSH

• Klíčová slova
• CRYPTORCHISM *, ESTROGENS *, KLINEFELTER'S SYNDROME *, SEMEN *,
• MeSH
• estrogeny * MeSH
• Klinefelterův syndrom * MeSH
• kontraceptiva ženská * MeSH
• kryptorchismus * MeSH
• lidé MeSH
• sperma * MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• estrogeny * MeSH
• kontraceptiva ženská * MeSH

Stagnant water bodies have generally received little attention regarding the presence of endocrine disruptive compounds, although they can integrate diverse pollutants from multiple different sources. Many compounds of anthropogenic as well as natural origin can contribute to the overall estrogenicity of surface waters and some of them can exhibit adverse effects on aquatic biota even in very low concentrations. This study focused on freshwater ponds and reservoirs affected by water blooms and determined the estrogenic activity of water by in vitro bioassay as well as concentrations of several important groups of estrogenic compounds (estrogenic hormones, alkylphenols, and phytoestrogens) by LC-MS/MS analyses. Estrogenic hormones were found at concentrations up to 7.1 ng.L-1, similarly to flavonoids, whose concentrations did not exceed 12.5 ng.L-1. Among alkylphenols, only bisphenol A and 4-tert-octylphenol were detected in levels reaching 100 ng.L-1 at maximum. Estrogenic activity of water samples varied from below the quantification limit to 1.95 ng.L-1. There does not seem to be any general causal link of the massive phytoplankton occurrence with the estrogenicity of water or concentration of phytoestrogens, since they showed no direct relationship with the phytoplankton abundance or composition across sites. The contribution of the analysed compounds to the estrogenic activity was calculated in three scenarios. In minimum scenario, just the compounds above quantification limit (LOQ) were taken into account and for most samples, only minor part (<6%) of the biological activity could be explained. In the mean and maximum scenarios, we included also compounds below LOQ into the calculations at the level of LOQ/2 and LOQ, respectively. In these cases, a considerable part of the estrogenic activity could be attributed to the possible presence of steroid estrogens below LOQ. However, for the samples with estrogenic activity greater than 1 ng.L-1, more than 50% of the estrogenic activity remained unexplained even in the maximum scenario. Probably other compounds or possible interactions between individual substances cause the estrogenic activity in these types of water bodies and in this case, the results of LC-MS/MS analyses cannot sufficiently predict the biological effects. A complex approach including bioassays is needed when assessing the estrogenicity of these types of surface waters.

• Klíčová slova
• Alkylphenols, Estrogenicity, Estrogens, Flavonoids, Surface water,
• MeSH
• chemické látky znečišťující vodu analýza   metabolismus   MeSH
• chromatografie kapalinová MeSH
• endokrinní disruptory analýza   metabolismus   MeSH
• estrogeny analýza   metabolismus   MeSH
• fytoplankton chemie   metabolismus   MeSH
• sladká voda chemie   MeSH
• tandemová hmotnostní spektrometrie MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chemické látky znečišťující vodu MeSH
• endokrinní disruptory MeSH
• estrogeny MeSH

Here is presented some of the first information on interactions of compounds produced by cyanobacteria and green algae with estrogen receptor signaling. Estrogenic potency of aqueous extracts and exudates (culture spent media with extracellular products) of seven species of cyanobacteria (10 different laboratory strains) and two algal species were assessed by use of in vitro trans-activation assays. Compounds produced by cyanobacteria and algae, and in particular those excreted from the cells, were estrogenic. Most exudates were estrogenic with potencies expressed at 50% of the maximum response under control of the estrogen receptor ranging from 0.2 to 7.2 ng 17β-estradiol (E(2)) equivalents (EEQ)/L. The greatest estrogenic potency was observed for exudates of Microcystis aerigunosa, a common species that forms water blooms. Aqueous extracts of both green algae, but only one species of cyanobacteria (Aphanizomenon gracile) elicited significant estrogenicity with EEQ ranging from 15 to 280 ng 17β-estradiol (E(2))/g dry weight. Scenedesmus quadricauda exudates and extracts of Aphanizomenon flos-aquae were antagonistic to the ER when coexposed to E(2). The EEQ potency was not correlated with concentrations of cyanotoxins, such as microcystin and cylindrospermopsin, which suggests that the EEQ was comprised of other compounds. The study demonstrates some differences between the estrogenic potency of aqueous extracts prepared from the same species, but of different origin, while the effects of exudates were comparable within species. The observed estrogenic potencies are important namely in relation to the possible mass expansion of cyanobacteria and release of the active compounds into surrounding water.

• MeSH
• Aphanizomenon metabolismus   MeSH
• biotest MeSH
• chemické látky znečišťující vodu metabolismus   farmakologie   MeSH
• Chlorophyta metabolismus   MeSH
• endokrinní disruptory metabolismus   farmakologie   MeSH
• estradiol metabolismus   farmakologie   MeSH
• estrogeny metabolismus   farmakologie   MeSH
• eutrofizace MeSH
• exsudáty a transsudáty chemie   MeSH
• mezibuněčná komunikace fyziologie   MeSH
• Microcystis účinky léků   MeSH
• receptory pro estrogeny metabolismus   MeSH
• sinice metabolismus   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chemické látky znečišťující vodu MeSH
• endokrinní disruptory MeSH
• estradiol MeSH
• estrogeny MeSH
• receptory pro estrogeny MeSH

The in vitro estrogen receptor (ER) reporter gene assay has long been used to measure estrogenic activity in wastewater. In a previous study, we demonstrated that the assay represents net estrogenic activity in the balance between estrogenic and antiestrogenic activities in wastewater. However, it remained unclear whether the net estrogenic activity measured by the in vitro ERα reporter gene assay can predict the in vivo estrogenic effect of wastewater. To determine this, we measured the following: estrogenic and antiestrogenic activities of wastewater and reclaimed water by the in vitro ERα reporter gene assay, expression of vitellogenin-1 (vtg1) and choriogenin-H (chgH) in male medaka (Oryzias latipes) by quantitative real-time PCR, and estrone, 17β-estradiol, estriol, and 17α-ethynylestradiol concentrations chemically to predict estrogenic activity. The net estrogenic activity measured by the in vitro medaka ERα reporter gene assay predicted the in vivo vtg1/chgH expression in male medaka more accurately than the concentrations of estrogens. These results also mean that in vivo vtg1/chgH expression in male medaka is determined by the balance between estrogenic and antiestrogenic activities. The in vitro medaka ERα reporter gene assay also predicted in vivo vtg1/chgH expression on male medaka better than the human ERα reporter gene assay.

• MeSH
• alfa receptor estrogenů analýza   genetika   metabolismus   MeSH
• biotest metody   MeSH
• chemické látky znečišťující vodu analýza   chemie   toxicita   MeSH
• estrogeny analýza   chemie   toxicita   MeSH
• exprese genu účinky léků   MeSH
• Oryzias MeSH
• vitelogeniny analýza   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• alfa receptor estrogenů MeSH
• chemické látky znečišťující vodu MeSH
• estrogeny MeSH
• vitelogeniny MeSH

Extracts of sediments from rivers in an industrialized area in the Czech Republic were used to evaluate suitability of a simple in vitro bioassay system to detect estrogen receptor (ER)-mediated activity in the complex mixture. Total estrogenic activity was detected by measuring luciferase activity in a stably transfected cell line containing an estrogen-responsive element linked to a luciferase reporter gene. For appropriate interpretation of ER-mediated activity, the effect of sediment extracts on the cell cytotoxicity was assessed at the same time. All sediment samples elicited considerable estrogenic activity. Fractionation of the extracts along with bioassay testing and subsequent instrumental analysis allowed the estrogenic fractions to be identified. The Florisil fraction, which was intermediate in polarity, was the most estrogenic. Instrumental analysis documented that the concentration of the degradation products of alkylphenol ethoxylates did not occur at sufficient concentrations to account for the estrogenic activity. Mass-balance calculations and testing of fractions confirmed that certain polycyclic aromatic hydrocarbons (PAHs) or their metabolites were the most likely compounds contributing to estrogenicity. Some other compounds, such as PCNs and PAH derivatives, that were present in the first and second fraction were tested for their potential estrogenic activity. Their ER-mediated activity and contribution to the overall responses of the complex extracts were very low. The concentrations of 17beta-estradiol present in the bioassay media was an important factor for the evaluation of (anti)estrogenicity of single compound(s) or complex mixtures.

• MeSH
• biotest MeSH
• estradiol MeSH
• estrogeny nesteroidní škodlivé účinky   MeSH
• geologické sedimenty chemie   MeSH
• látky znečišťující životní prostředí škodlivé účinky   MeSH
• lidé MeSH
• monitorování životního prostředí MeSH
• nádorové buňky kultivované MeSH
• nádory prsu patologie   MeSH
• polycyklické aromatické uhlovodíky škodlivé účinky   MeSH
• receptory pro estrogeny účinky léků   fyziologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• estradiol MeSH
• estrogeny nesteroidní MeSH
• látky znečišťující životní prostředí MeSH
• polycyklické aromatické uhlovodíky MeSH
• receptory pro estrogeny MeSH

This review discusses the potential contribution of phytoestrogens and mycoestrogens to in vitro estrogenic activities occurring in surface waters and in vivo estrogenic effects in fish. Main types, sources, and pathways of entry into aquatic environment of these detected compounds were summarized. Reviewed concentrations of phyto/mycoestrogens in surface waters were mostly undetectable or in low ng/L ranges, but exceeded tens of μg/L for the flavonoids biochanin A, daidzein and genistein at some sites. While a few phytosterols were reported to occur at relatively high concentrations in surface waters, information about their potencies in in vitro systems is very limited, and contradictory in some cases. The relative estrogenic activities of compounds (compared to standard estrogen 17β-estradiol) by various in vitro assays were included, and found to differ by orders of magnitude. These potencies were used to estimate total potential estrogenic activities based on chemical analyses of phyto/mycoestrogens. In vivo effective concentrations of waterborne phyto/mycoestrogens were available only for biochanin A, daidzein, formononetin, genistein, equol, sitosterol, and zearalenone. The lowest observable effect concentrations in vivo were reported for the mycoestrogen zearalenone. This compound and especially its metabolites also elicited the highest in vitro estrogenic potencies. Despite the limited information available, the review documents low contribution of phyto/mycoestrogens to estrogenic activity in vast majority of surface waters, but significant contribution to in vitro responses and potentially also to in vivo effects in areas with high concentrations.

• Klíčová slova
• Estrogenic equivalent, Estrogenicity, Flavonoids, Mycoestrogen, Phytoestrogen, Phytosterols, Relative potency, Sterol, Surface waters,
• MeSH
• chemické látky znečišťující vodu * analýza   toxicita   MeSH
• fytoestrogeny * analýza   toxicita   MeSH
• isoflavony * analýza   toxicita   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• chemické látky znečišťující vodu * MeSH
• fytoestrogeny * MeSH
• isoflavony * MeSH

Aquatic biotests are important tools targeting various effects in ecotoxicology, including endocrine disruption. Unintentional exposure of bioassay organisms to endocrine disruptors during cultivation or testing may interfere with assessed endpoints. We illustrate this issue on the example of laboratory phytoplankton cultivation, where possible sources of estrogenic compounds have been revealed. Fifty-four blank samples (water and fresh or cultivated growth media) were assessed by in vitro biotests for their estrogenicity, and major known estrogens originating from plastic materials, bisphenol A and alkylphenols, were analyzed in selected samples. The samples of freshly prepared growth medium elicited weak estrogenic response in bioassays and some samples of the aerated media caused responses even above the 50% of maximum of the reference compound (17β-estradiol, E2), while the samples from diverse laboratory water sources did not show significant estrogenic activity. The results identified substances contained in the growth medium as minor but reproducible contributors to estrogenicity in the cultivations. Sporadic but significant effects (up to 4.9 ng E2 equivalent/L) can be ascribed to compounds released from the used plastic materials during aeration of the cultivations. The potential sources of unintentional exposure to estrogenic compounds need to be considered in aquatic cultivations and biotests, since they could impact their outcomes, especially in arrangements assessing reproduction or whole life cycle biotests, or production of bioactive compounds by phytoplankton. The findings emphasize the necessity to assess all relevant blanks, ideally by sensitive high throughput in vitro assays that reflect also unknown pollutants and minimize all potential sources of background contamination. In vitro assays show very good applicability for this purpose since they enable to screen for any background estrogenicity of the used media and materials without the need of analyzing individual compounds, which often might not be known.

• Klíčová slova
• Aeration, Alkylphenols, Aquatic medium, Bisphenol A, Estrogenicity,
• MeSH
• chemické látky znečišťující vodu * toxicita   MeSH
• endokrinní disruptory * analýza   toxicita   MeSH
• estrogeny analýza   toxicita   MeSH
• fytoplankton MeSH
• monitorování životního prostředí metody   MeSH
• plastické hmoty MeSH
• voda MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• chemické látky znečišťující vodu * MeSH
• endokrinní disruptory * MeSH
• estrogeny MeSH
• plastické hmoty MeSH
• voda MeSH