Ionic liquid-functionalized covalent organic frameworks on the surface of silica for online solid-phase extraction.

A covalent organic framework (COF) was gown on porous silica with 1,3,5-tri(4-aminophenyl)benzene and 2,5-divinyl-1,4-phenyldiformaldehyde as monomers, and two ionic liquids were grafted to COF by a click reaction. The materials before and after the modification of ionic liquids were separately packed into solid-phase extraction columns (10 × 4.6 mm, i.d.), which were coupled with liquid chromatography to construct online analysis systems. The extraction mechanisms of polycyclic aromatic hydrocarbons, bisphenols, diphenylalkanes and benzoic acids were investigated on these materials. There were π-π, hydrogen-bond and electrostatic interactions on ionic liquid-functionalized sorbents. After the comparison among these materials, the best sorbent was used, and the analytical method was established and successfully applied to the detection of some estrogens from actual samples. For the analytical method, the detection limit was as low as 0.005 µg L-1, linear range was as wide as 0.017-10.0 µg L-1, and enrichment ratio was as high as 3635. The recoveries in actual samples were 70 %-129 %.

Aquatic toxicity, bioaccumulation potential, and human estrogen/androgen activity of three oxo-Liquid Organic Hydrogen Carrier (oxo-LOHC) systems.

The Liquid Organic Hydrogen Carrier (LOHC) technology offers a technically attractive way for hydrogen storage. If LOHC systems were to fully replace liquid fossil fuels, they would need to be handled at the multi-million tonne scale. To date, LOHC systems on the market based on toluene or benzyltoluene still offer potential for improvements. Thus, it is of great interest to investigate potential LOHCs that promise better performance and environmental/human hazard profiles. In this context, we investigated the acute aquatic toxicity of oxygen-containing LOHC (oxo-LOHC) systems. Toxic Ratio (TR) values of oxo-LOHC compounds classify them baseline toxicants (0.1 < TR < 10). Additionally, the mixture toxicity test conducted with D. magna suggests that the overall toxicity of a benzophenone-based system can be accurately predicted using a concentration addition model. The estimation of bioconcentration factors (BCF) through the use of the membrane-water partition coefficient indicates that oxo-LOHCs are unlikely to be bioaccumulative (BCF < 2000). None of the oxo-LOHC compounds exhibited hormonal disrupting activities at the tested concentration of 2 mg/L in yeast-based reporter gene assays. Therefore, the oxo-LOHC systems seem to pose a low level of hazard and deserve more attention in ongoing studies searching for the best hydrogen storage technologies.

Preparation of deep eutectic solvent modified magnetic graphene oxide/metal organic framework nanocomposites for the extraction of three estrogens in cosmetics.

A novel magnetic dispersive solid phase extraction (MDSPE) procedure based on the deep eutectic solvent (DES) modified magnetic graphene oxide/metal organic frameworks nanocomposites (MGO@ZIF-8@DES) was established and used for the efficient enrichment of estradiol, estrone, and diethylstilbestrol in cosmetics (toner, lotion, and cream) for the first time. Then, the three estrogens were separated and determined by UHPLC-UV analysis method. In order to study the features and morphology of the synthesized adsorbents, various techniques such as FT-IR, SEM, and VSM measurements were executed. The MGO@ZIF-8@DES nanocomposites combine the advantages of high adsorption capacity, adequate stability in aqueous solution, and convenient separation from the sample solution. To achieve high extraction recoveries, the Box-Behnken design and single factor experiment were applied in the experimental design. Under the optimum conditions, the method detection limits for three estrogens were 20-30 ng g-1. This approach showed a good correlation coefficient (r more than 0.9998) and reasonable linearity in the range 70-10000 ng g-1. The relative standard deviations for intra-day and inter-day were beneath 7.5% and 8.9%, respectively. The developed MDSPE-UHPLC-UV method was successfully used to determine  three estrogens in cosmetics, and acceptable recoveries in the intervals of 83.5-95.9% were obtained. Finally, three estrogens were not detected in some cosmetic samples. In addition, the Complex GAPI tool was used to evaluate the greenness of the developed pretreatment method. The developed MDSPE-UHPLC-UV method is sensitive, accurate, rapid, and eco-friendly, which provides a promising strategy for determining hormones in different complex samples.

Reduced estrogenic risks of a sunscreen additive: Theoretical design and evaluation of functionally improved salicylates.

Salicylic esters (SEs), the widely used ultraviolet (UV) absorbers in sunscreen products, have been found to have health risks such as skin sensitization and estrogenic effects. This study aims to design SE substitutes that maintain high UV absorbance while reducing estrogenicity. Using molecular docking and Gaussian09 software for initial assessments and further application of a combination of two-dimensional and three-dimensional quantitative structure-activity relationships (2D-QSAR and 3D-QSAR, respectively) models, we designed 73 substitutes. The best-performing molecules, ethylhexyl salicylate (EHS)-5 and EHS-15, significantly reduced estrogenicity (44.54 % and 17.60 %, respectively) and enhanced UV absorbance (249.56 % and 46.94 %, respectively). Through screening for human health risks, we found that EHS-5 and EHS-15 were free from skin sensitivity and eye irritation and exhibited reduced skin permeability compared with EHS. Furthermore, the photolysis and synthetic pathways of EHS-5 and EHS-15 were deduced, demonstrating their good photodegradability and potential synthesizability. In addition, we analyzed the mechanisms underlying the changes in estrogenic effects and UV absorption properties. We identified covalent hydrogen bond basicity and acidity Propgen value for atomic molecular properties and the highest occupied molecular orbital eigenvalue as the main factors affecting the estrogenic effect and UV absorbance of SEs, respectively. This study focuses on the design and screening of SEs, exhibiting enhanced functionality, reduced health risks, and synthetic feasibility.

Direct extraction of ten estrogens from milk samples with DVB/NVP-modified magnetic solid-phase extraction adsorbent followed by pre-column derivatization-UHPLC-MS/MS.

Estrogens and their analogues can cause harm to human health through the food chain. Ten estrogens in different milk samples were directly extracted by amphiphilic divinylbenzene/N-vinyl-2-pyrrolidone (DVB/NVP)-Fe3O4@SiO2-based magnetic solid-phase extraction (MSPE) followed by pre-column derivatization and ultra-high performance liquid chromatography tandem mass-spectrometry (UHPLC-MS/MS) detection. Under the optimal conditions, the limits of detection for ten analytes were in the range of 0.05-0.38 ng mL-1 in whole liquid milk matrix and 0.04-3.00 ng g-1 in milk powder matrix. The intra-/inter-day accuracy ranged in 83.4-113.8%, with RSDs in 2.5-15.0%. A total of 15 brands of liquid milk and milk powder samples were analyzed, and only estradiol was detected in three brands of boxed liquid milk within safe range. The proposed sample pretreatment eliminated the common protein precipitation process, improved the sample throughput, and has the potential for routine testing of estrogens and their analogues in market-sale milk samples.

Lignin-derivable alternatives to bisphenol A with potentially undetectable estrogenic activity and minimal developmental toxicity.

Lignin-derivable bisguaiacols/bissyringols are viable alternatives to commercial bisphenols; however, many bisguaiacols/bissyringols (e.g., bisguaiacol F [BGF]) have unsubstituted bridging carbons between the aromatic rings, making them more structurally similar to bisphenol F (BPF) than bisphenol A (BPA) - both of which are suspected endocrine disruptors. Herein, we investigated the estrogenic activity (EA) and developmental toxicity of dimethyl-substituted bridging carbon-based lignin-derivable bisphenols (bisguaiacol A [BGA] and bissyringol A [BSA]). Notably, BSA showed undetectable EA at seven test concentrations (from 10-12 M to 10-6 M) in the MCF-7 cell proliferation assay, whereas BPA had detectable EA at five concentrations (from 10-10 M to 10-6 M). In silico results indicated that BSA had the lowest binding affinity with estrogen receptors. Moreover, in vivo chicken embryonic assay results revealed that lignin-derivable monomers had minimal developmental toxicity vs. BPA at environmentally relevant test concentrations (8.7-116 µg/kg). Additionally, all lignin-derivable compounds showed significantly lower expression fold changes (from ∼1.81 to ∼4.41) in chicken fetal liver tests for an estrogen-response gene (apolipoprotein II) in comparison to BPA (fold change of ∼11.51), which was indicative of significantly reduced estrogenic response. Altogether, the methoxy substituents on lignin-derivable bisphenols appeared to be a positive factor in reducing the EA of BPA alternatives.

Asymmetric allostery in estrogen receptor-α homodimers drives responses to the ensemble of estrogens in the hormonal milieu.

The estrogen receptor-α (ER) is thought to function only as a homodimer but responds to a variety of environmental, metazoan, and therapeutic estrogens at subsaturating doses, supporting binding mixtures of ligands as well as dimers that are only partially occupied. Here, we present a series of flexible ER ligands that bind to receptor dimers with individual ligand poses favoring distinct receptor conformations-receptor conformational heterodimers-mimicking the binding of two different ligands. Molecular dynamics simulations showed that the pairs of different ligand poses changed the correlated motion across the dimer interface to generate asymmetric communication between the dimer interface, the ligands, and the surface binding sites for epigenetic regulatory proteins. By examining the binding of the same ligand in crystal structures of ER in the agonist vs. antagonist conformers, we also showed that these allosteric signals are bidirectional. The receptor conformer can drive different ligand binding modes to support agonist vs. antagonist activity profiles, a revision of ligand binding theory that has focused on unidirectional signaling from the ligand to the coregulator binding site. We also observed differences in the allosteric signals between ligand and coregulator binding sites in the monomeric vs. dimeric receptor, and when bound by two different ligands, states that are physiologically relevant. Thus, ER conformational heterodimers integrate two different ligand-regulated activity profiles, representing different modes for ligand-dependent regulation of ER activity.

Solvent and monomer regulation synthesis of core-shelled magnetic ß-cyclodextrin microporous organic network for efficient extraction of estrogens in biological samples prior to HPLC analysis.

The abnormal estrogens levels in human body can cause many side effects and diseases, but the quantitative detection of the trace estrogens in complex biological samples still remains great challenge. Here we reported the fabrication of a novel core-shell structured magnetic cyclodextrin microporous organic network (Fe3O4@CD-MON) for rapid magnetic solid phase extraction (MSPE) of four estrogens in human serum and urine samples prior to HPLC-UV determination. The uniform spherical core-shell Fe3O4@CD-MONs was successfully regulated by altering the reactive monomers and solvents. The Fe3O4@CD-MONs owned high specific surface area, good hydrophobicity, large superparamagnetism, and abundant extraction sites for estrogens. Under optimal conditions, the proposed MSPE-HPLC-UV method provided wide linearity range (2.0-400 µg L-1), low limits of detection (0.5-1.0 µg L-1), large enrichment factors (183-198), less adsorbent consumption (3 mg), short extraction time (3 min), and good stability and reusability (at least 8 cycles). The established method had also been successfully applied to the enrichment and detection of four estrogens in serum and urine samples with a recovery of 88.4-105.1 % and a relative standard deviation of 1.0-5.9 %. This work confirmed the feasibility of solvent and monomer regulation synthesis of Fe3O4@CD-MON composites, and revealed the great prospects of magnetic CD-MONs for efficient enrichment of trace estrogens in complex biological samples.

Synthesis and estrogenic activity of BODIPY-labeled estradiol conjugates.

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)-17ß-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.

Degradation and humification of steroidal estrogens in the soil environment: A review.

Emerging pollutants are toxic and harmful chemical substances characterized by environmental persistence, bioaccumulation and biotoxicity, which can harm the ecological environment and even threaten human health. There are four categories of emerging pollutants that are causing widespread concern, namely, persistent organic pollutants, endocrine disruptors, antibiotics, and microplastics. The distribution of emerging pollutants has spatial and temporal heterogeneity, which is influenced by factors such as geographical location, climatic conditions, population density, emission amount, etc. Steroidal estrogens (SEs) discussed in this paper belong to the category of endocrine disruptors. There are generally three types of fate for SEs in the soil environment: sorption, degradation and humification. Humification is a promising pathway for the removal of SEs, especially for those that are difficult to degrade. Through humification, these difficult-to-degrade SEs can be effectively transferred or fixed, thus reducing their impact on the environment and organisms. Contrary to the well-studied process of sorption and degradation, the role and promise of the humification process for the removal of SEs has been underestimated. Based on the existing research, this paper reviews the sources, classification, properties, hazards and environmental behaviors of SEs in soil, and focuses on the degradation and humification processes of SEs and the environmental factors affecting their processes, such as temperature, pH, etc. It aims to provide references for the follow-up research of SEs, and advocates further research on the humification of organic pollutants in future studies.

Assessment of estrogenic potential from exudates of microcystin-producing and non-microcystin-producing Microcystis by metabolomics, machine learning and E-screen assay.

Cyanobacterial blooms, often dominated by Microcystis aeruginosa, are capable of producing estrogenic effects. It is important to identify specific estrogenic compounds produced by cyanobacteria, though this can prove challenging owing to the complexity of exudate mixtures. In this study, we used untargeted metabolomics to compare components of exudates from microcystin-producing and non-microcystin-producing M. aeruginosa strains that differed with respect to their ability to produce microcystins, and across two growth phases. We identified 416 chemicals and found that the two strains produced similar components, mainly organoheterocyclic compounds (20.2%), organic acids and derivatives (17.3%), phenylpropanoids and polyketides (12.7%), benzenoids (12.0%), lipids and lipid-like molecules (11.5%), and organic oxygen compounds (10.1%). We then predicted estrogenic compounds from this group using random forest machine learning. Six compounds (daidzin, biochanin A, phenylethylamine, rhein, o-Cresol, and arbutin) belonging to phenylpropanoids and polyketides (3), benzenoids (2), and organic oxygen compound (1) were tested and exhibited estrogenic potency based upon the E-screen assay. This study confirmed that both Microcystis strains produce exudates that contain compounds with estrogenic properties, a growing concern in cyanobacteria management.

Deciphering the synergistic effects of photolysis and biofiltration to actuate elimination of estrogens in natural water matrix.

The presence of estrogens in water environments has raised concerns for human health and ecosystems balance. These substances possess potent estrogenic properties, causing severe disruptions in endocrine systems and leading to reproductive and developmental problems. Unfortunately, conventional treatment methods struggle to effectively remove estrogens and mitigate their effects, necessitating technological innovation. This study investigates the effectiveness of a novel sequential photolysis-granular activated carbon (GAC) sandwich biofiltration (GSBF) system in removing estrogens (E1, E2, E3, and EE2) and improving general water quality parameters. The results indicate that combining photolysis pre-treatment with GSBF consistently achieved satisfactory performance in terms of turbidity, dissolved organic carbon (DOC), UV254, and microbial reduction, with over 77.5 %, 80.2 %, 89.7 %, and 92 % reduction, respectively. Furthermore, this approach effectively controlled the growth of microbial biomass under UV irradiation, preventing excessive head loss. To assess estrogen removal, liquid chromatography-tandem mass spectrometry (LC-MS) measured their concentrations, while bioassays determined estrogenicity. The findings demonstrate that GSBF systems, with and without photolysis installation, achieved over 96.2 % removal for estrogens when the spike concentration of each targeted compound was 10 µg L-1, successfully reducing estrogenicity (EA/EA0) to levels below 0.05. Additionally, the study evaluated the impact of different thicknesses of GAC layer filling (8 cm, 16 cm, and 24 cm) and found no significant difference (p>0.05) in estrogen and estrogenicity removal among them.

A 4-year study of bovine reproductive hormones that are induced by pharmaceuticals and appear as steroid estrogenic pollutants in the resulting slurry, using in vitro and instrumental analytical methods.

The main objective of the research was to study the environmental "price" of the large-scale, milk production from a rarely known perspective, from the mapping of the estrogenic footprint (the amount of oestrus-inducer hormonal products, and the generated endoestrogens) in the resulting slurry in a dairy cow farm. These micropollutants are endocrine-disrupting chemicals (EDCs) and can be dangerous to the normal reproductive functions even at ng/kg concentration. One of them, 17ß-estradiol, has a 20,000 times stronger estrogenic effect than bisphenol-A, a widely known EDC of industrial origin. While most studies on EDCs are short-term and/or laboratory based, this study is longitudinal and field-based. We sampled the slurry pool on a quarterly basis between 2017 and 2020. Our purpose was testing the estrogenic effects using a dual approach. As an effect-based, holistic method, we developed and used the YES (yeast estrogen screen) test employing the genetically modified Saccharomyces cerevisiae BJ3505 strain which contains human estrogenic receptor. For testing exact molecules, UHPLC-FLD was used. Our study points out that slurry contains a growing amount of EDCs with the risk of penetrating into the soil, crops and the food chain. Considering the Green Chemistry concept, the most benign ways to prevent of the pollution of the slurry is choosing appropriate oestrus-inducing veterinary pharmaceuticals (OIVPs) and the separation of the solid and liquid parts with adequate treatment methods. To our knowledge, this is the first paper on the adaptation of the YES test for medicine and slurry samples, extending its applicability. The adapted YES test turned out to be a sensitive, robust and reliable method for testing samples with potential estrogenic effect. Our dual approach was successful in evaluating the estrogenic effect of the slurry samples.

[Sorption Characteristics and Site Energy Distribution Theory of Typical Estrogens on Microplastics].

Microplastics (MPs) and estrogens are high-profile emerging contaminants at present, and MPs might become the carrier of estrogens in the environment and induce combined pollution. To study the adsorption behavior of polyethylene (PE) microplastics to typical estrogens, the adsorption isothermal properties of the six estrogens[estrone (E1), 17α-estradiol (17α-E2), 17ß-estradiol (17ß-E2), estriol (E3), diethylstilbestrol (DES), and ethinylestradiol (17α-EE2)] in single-solute and mixed-solute systems were studied through batch equilibrium adsorption experiments, in which the PE microplastics before and after adsorption were characterized by X-ray photoelectron spectroscopy (XPS) and Fourier transform infrared spectroscopy (FTIR). Then, the site energy distribution theory of the adsorption of six estrogens on PE microplastics was further analyzed based on the Freundlich model. The results showed that the adsorption process of selected estrogens with two concentrations (100 µg·L-1 and 1000 µg·L-1) on PE were more consistent with the pseudo-second order kinetic model. The increase in initial concentration reduced the equilibrium time of adsorption and increased the adsorbing capacity of estrogens on PE. In the single system (one estrogen) or mixed system (six estrogens) with different concentrations (10 µg·L-1-2000 µg·L-1), the Freundlich model showed the best fitting effect for the adsorption isotherm data (R2>0.94). The results of isothermal adsorption experiments and XPS and FTIR spectra showed that the adsorption of estrogens on PE in the two systems was heterogeneous adsorption, and hydrophobic distribution and van der Waals forces were the principal factors in the process of adsorption. The occurrence of C-O-C (in only the DES and 17α-EE2 systems) and O-C[FY=,1]O (in only the 17α-EE2 system) indicated that the adsorption of synthetic estrogens on PE was affected slightly by chemical bonding function, but no obvious effects were observed for natural estrogens. The results of site energy distribution analysis showed that, compared with the single system, the adsorption site energy of each estrogen shifted to the high-energy region in its entirety in the mixed system, and the site energy increased by 2.15%-40.98%. The energy change in DES was the most significant among all of the estrogens, indicating its competitive advantage in the mixed system. The above results of this study can provide some reference for the study of adsorption behavior, mechanism of action, and environmental risks under the coexisting condition of organic pollutants and MPs.

Synthetic ERRα/ß/γ Agonist Induces an ERRα-Dependent Acute Aerobic Exercise Response and Enhances Exercise Capacity.

Repetitive physical exercise induces physiological adaptations in skeletal muscle that improves exercise performance and is effective for the prevention and treatment of several diseases. Genetic evidence indicates that the orphan nuclear receptors estrogen receptor-related receptors (ERRs) play an important role in skeletal muscle exercise capacity. Three ERR subtypes exist (ERRα, ß, and γ), and although ERRß/γ agonists have been designed, there have been significant difficulties in designing compounds with ERRα agonist activity. Additionally, there are limited synthetic agonists that can be used to target ERRs in vivo. Here, we report the identification of a synthetic ERR pan agonist, SLU-PP-332, that targets all three ERRs but has the highest potency for ERRα. Additionally, SLU-PP-332 has sufficient pharmacokinetic properties to be used as an in vivo chemical tool. SLU-PP-332 increases mitochondrial function and cellular respiration in a skeletal muscle cell line. When administered to mice, SLU-PP-332 increased the type IIa oxidative skeletal muscle fibers and enhanced exercise endurance. We also observed that SLU-PP-332 induced an ERRα-specific acute aerobic exercise genetic program, and the ERRα activation was critical for enhancing exercise endurance in mice. These data indicate the feasibility of targeting ERRα for the development of compounds that act as exercise mimetics that may be effective in the treatment of numerous metabolic disorders and to improve muscle function in the aging.

Rapid Detection of Estrogens in Cosmetics by Chemical Derivatization and Paper-Spray Ionization Mass-Spectrometry.

Estrogens in personal care products are harmful to customers. Conventional methods such as HPLC and LC-MS require tedious sample pretreatment and long analytical time. Paper-spray ionization mass spectrometry (PSI-MS) is a powerful tool for the determination of compounds with little time and minimal pretreatment procedures. Since most estrogens show poor responses in PSI-MS, we developed a chemical derivatization and PSI-MS method to determinate three estrogens: estradiol, estriol and ethinyloestradiol with estradiol valerate as the internal standard (I.S.). After derivatization with 2-fluoro-1-methyl-pyridinium-p-toluene-sulfonate, the three estrogens could be quantified in seconds. This method showed good linearity in the range of 0.1~30 µg·mL-1, with R2 > 0.999. Their recovery results were all between 85%~115%. The limits of detection (LOD) were 0.04 µg·mL-1, 0.02 µg·mL-1 and 0.02 µg·mL-1 for estradiol, estriol and ethinyloestradiol respectively, which improved around 200, 2000, and 900 times compared to non-derivative PSI-MS. The method could quantitatively determine estrogens in cosmetics.

Interferences in the yeast estrogen screen (YES) assay for evaluation of estrogenicity in environmental samples, chemical mixtures, and individual substances.

The Yeast Estrogen Screen (YES) has a specific mechanism of action that allows for the analysis of estrogenic EDC at low concentrations, and it has been broadly used to estimate the estrogenic potential of environmental samples. However, the experimental parameters of this assay still demand an investigation, such as cell density, incubation time, wavelength on the experimental outcome, cytotoxicity, and estrogenic activity adsorbed on suspended solids. We studied these interferences and applied the assay to single substances, mixtures, and environmental matrices from different sources. The increase in cell density amplifies the assay sensitivity only to a limited extent, while the reduction in incubation time decreased assay sensitivity - although it was not significant for surface water, no differences were observed between estradiol-equivalents derived of 48 h and 72 h measurements. The particulate phase was of utmost importance for the total estrogenic activity of the landfill leachate and surface water. Surface waters, landfill leachates and sediments also showed antiestrogenic activity and the integration of both estrogenic and antiestrogenic endpoints provided deeper insights into the potential risk associated with EDC. This study elucidated experimental interferences that may arise during the implementation and use of this assay, bringing more understanding to experimental parameters during the application of the assay for estrogenicity screening.

Establishment and validation of a sensitive LC-MS/MS method for quantification of urinary estrogens in women with gestational diabetes mellitus.

Gestational diabetes mellitus (GDM) is not only a threat to the health of pregnant women, but also has profound effects on the health of offspring. Studies have shown that the imbalance of estrogen metabolism is associated with an increased risk of GDM. In this study, an ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was established and validated for simultaneous quantification of thirteen estrogens in the urine of GDM women, including estrone (E1), estradiol (E2), estriol (E3), and their hydroxylated and methylated metabolites. The method was achieved on a Waters CORTECS C18 column (2.1 mm × 150 mm, 1.6 µm) within 8.5 min. The linear range of thirteen estrogens in urine was 2-1000 pg·mL-1. Both intra- and inter-day precision for each analyte were less than 15%, with accuracies ranging from 8.3% to 7.3%. The extraction recoveries rate were between 86% and 111%, and stability verification results met the requirements for determination of biological samples. The results suggested that the concentrations of estrogens in all urine samples range from 0.08 to 134.06 (pg·mg-1 creatinine). The mean levels of E1, E2 and most estrogen metabolites in the urine of GDM women were higher than those in healthy pregnant women. Notably, the mean level of 2-hydroxyestrone (2-OHE1) in GDM women was 13.2-fold lower than that in healthy pregnant women. The types of estrogens with the highest mean levels in the urine of GDM and healthy pregnant women were obviously different, which are 2-methoxyestrone (2MeOE1) and E3, respectively. Our results demonstrated that this specific and sensitive method is suitable for quantifying estrogens in human urine and could provide support for further research on estrogen-related pathological mechanisms in GDM and other diseases.

Determination of estrogens in human serum using a novel chemical derivatization-assisted liquid chromatography-electrospray ionization-tandem mass spectrometry method.

RATIONALE: Assessing estrogen concentrations in biological systems can provide valuable information on physiological processes, which is crucial for the early diagnosis of many diseases. Because estrogens are present in the human body in low concentrations and in a wide dynamic range, analytical methods with high sensitivity and specificity are required for their determination in complex biological matrices. METHODS: To discover an appropriate derivatization reagent for estrogen mass spectrometry (MS) analysis, we compared five sulfonyl chloride derivatization reagents, namely 3-methyl-8-quinolinesulfonyl chloride (MQSCl) and 8-quinolinesulfonyl chloride (QSCl), 1-methyl-1H-pyrazole-4-sulfonyl chloride, 1,2-methyl-imidazole-5-sulfonyl chloride, and dansyl chloride. By selecting the derivatization reagent with the best performance, we developed and validated a novel chemical derivatization-assisted-liquid chromatography-electrospray ionization-tandem mass spectrometry (CD-LC-ESI-MS/MS) method to simultaneously determine the concentrations of estrone, estradiol, and estriol (E1, E2, and E3) in human serum. RESULTS: It was found that among the five investigated reagents, MQSCl-derivatized estrogens presented the highest sensitivity using LC-ESI-MS/MS. Based on this discovery, MQSCl was chosen to derivatize the analyzed estrogens to assist LC-ESI-MS/MS analysis. The limit of quantification of E1, E2, and E3 was measured as 2.7, 4.6, and 5.1 pg/mL, respectively. Inter- and intra-day precision, expressed as the coefficient of variation, was shown to be lower than 13.2% for all concentrations. The mean recovery was 72.4% overall, with good reproducibility at low, medium, and high concentrations in the calibration range. CONCLUSIONS: The developed method was successfully applied to the quantitative determination of estrogens in clinical human serum from pediatric and adult women, demonstrating the suitability of estrogen analysis in the biological matrix at low concentration (pg/mL).

Novel Evidence-Based Combination of Plant Extracts with Multitarget Mechanisms of Action for the Elimination of Hot Flashes during Menopause.

Hot flashes are considered the most bothersome complaint during menopause. Although hormone therapy is an effective option to relieve hot flashes, it has been associated with significant side effects. The aim of our study is to suggest a novel combination of different plant extracts with distinct mechanisms of action against hot flashes. We selected the rhizome of Glycyrrhiza glabra L. (Fabaceae), the rhizome of Actaea racemosa L. (Ranunculaceae), the aerial parts of Hypericum perforatum L. (Hypericaceae) to produce extracts rich in bioactive phytochemicals and the seed oil of Oenothera biennis L. (Onagraceae). We investigated their estrogenic and antioxidant potential and their inhibitory effect against prostaglandin D2 receptor 1 (DP1) as a novel mechanistic pathway for vasodilation in hot flashes, alone or in combination. The phytochemical footprint of the extracts was analyzed using HPLC-PDA and UPLC-HRMS. We observed that the tested extracts possess different mechanisms of action. A. racemosa exerts a beneficial activation of the estrogen receptor, H. perforatum possesses the highest antioxidant capacity and the seed oil of O. biennis inhibits the DP1 receptor. The triple combination in the optimal doses pertains to efficacy against all three mechanisms of action, serves as a multitarget plant-based therapy and could serve as a novel strategy for the alleviation of hot flashes in postmenopausal women.