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Androstenedione, testosterone, estrone and estradiol concentrations and estrogen-to-androgen ratios in the ovaries of female rats treated with atrazine, vinclozolin, methoxychlor, and bisphenol A daily for 2 weeks. The figure represents: the mean ovarian steroid concentrations after a 2-week EDC treatment or after the recovery period following the 2-week EDC treatment and the ovarian estrone-to- androstenedione and estradiol- to-testosterone ratios (pg estro- gens/ng androgens) in female adult rats treated or not with EDCs for 2-week. The values are the mean±SEM of 5 animals per group. * p <0.05; ** p <0.01 versus the corresponding control (control 1 for atrazine treatment, control 2 for vinclozolin, methoxychlor, and bisphenol A treatment)
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Reproductive function is controlled by a finely tuned balance of androgens and estrogens. Environmental toxicants, notably endocrine disrupting chemicals (EDCs), appear to be involved in the disruption of hormonal balance in several studies. To further describe the effects of selected EDCs on steroid secretion in female rats, we aim to simultaneous...
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... analytical method developed in this study was applied to the rat ovaries obtained from the previously described subacute in vivo study. The results are summarized in Table 4 and Fig. 6. See Electronic supplementary material for examples of chromatograms. Table 4 shows the ovarian concentrations of ATZ, BPA, HPTE, and M2. After the recovery period, the chemicals could not be ...
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... steroid concentrations in the ovaries are presented in Fig. 6. After a 2-week treatment with ATZ, females showed slightly increased ovarian estrone. VCZ treatment significantly increased ovarian androstenedione and estrone. MXC-treated females showed a decrease in androstenedione and testosterone concentrations in the ovaries. BPA treatment showed no significant effect. After the recovery period, there was no more significant effect on sex steroid levels. As far as estrogen-to-androgen ratios are concerned, ATZ and MXC treatments significantly increased the ovarian estrone-to-androstenedione ratio. An over- or under-secretion of one or more steroids can cause a steroidal imbalance that results in various disorders. Precise steroid analysis is very important, not only to predict pathological alterations, but also to understand toxicity mechanisms. Indeed, small variations in steroid concentrations can induce major toxicological changes, particularly when exposure occurs during a critical win- dow [35]. There are many challenges when analyzing steroids, due to their low concentrations and their struc- tural similarities. The method developed herein aimed to simultaneously quantify sex steroids and EDCs in rat ovaries. The excellent recoveries obtained for the sex hormones (Table 2), as compared with the recoveries reported in the literature (higher than 80 %) [10], highlight the efficiency of the “ QuEChERS ” approach for EDCs as well as for hormone extraction. Moreover, the biases of the recoveries as determined by the RSDs are sufficient and acceptable because they are lower than 20 % (Table 2). As ovaries are complex matrices, it was important to thoroughly study the matrix effect aspect. Even if ion suppression was observed, the low variations of recovery, illustrated by low values of matrix effect (Table 2), show the efficiency of the extraction process. To the best of our knowledge, this is the first time that “ QuEChERS ” sample preparation was applied to rat ovaries with analyte concentrations reported in the nanogram range per gram of tissue and the first time that the simultaneous analysis of hormones and EDCs was conducted. LODs and LOQs are similar to those obtained with tissues from other species [10]. Furthermore, the method was sufficiently sensitive and precise to accurately quantify ovarian steroid and EDCs levels in female rats. To better characterize the effects of EDCs in female rats, we focused our work on hormone production in the ovaries after in vivo EDC exposure. We took into account not only the effect on absolute sex steroid concentration, but also on the estrogen-to-androgen ratio and the reproductive side effects. We discuss below the potential mechanisms for altered hormone balance. Following EDC treatment, some females presented estrous cycle disruption and histopatho- logical disorders of the uterus or ovaries (Table 1). These alterations were caused by hormonal balance disruption or altered receptor availability. Ovarian sex steroid concentrations were altered following ATZ, VCZ, and MXC treatments (Fig. 6). ATZ, MXC, and BPA are classified as estrogenic, according to in vitro studies [4, 5, 7]. However, the effects on hormonal balance in our study are more or less important. ATZ elevated the estrone-to-androstenedione ratio by increasing estrone. This effect was in accordance with several other studies, which associated ATZ with disturban- ces in the balance between androgens and estrogens in male rats [36]. This elevated estrogen-to-androgen ratio could be specifically linked to aromatase activity induction observed in in vitro studies [37]. Ovaries from females treated with MXC showed normal estrogen levels and decreased androgen levels. Alteration of steroidogenesis resulting in an androgen decrease has been previously described in male rats following MXC treatment [38]. Thus, MXC appears to exert an anti-androgenic activity in rats. As the effect of BPA on rat ovaries was not significant, we cannot conclude what effect BPA had on the sex steroid levels in our study. VCZ treatment has been associated with anti-androgenic effects in vitro, via the antagonistic binding of its metabolite M2 to the androgen receptor [6]. In our study, VCZ increased ovarian androstenedione and testosterone levels, an effect which is opposite to what is usually reported for this class of compound (anti-androgenic). This effect could account for a direct steroidogenic up-regulation, previously reported in rat and human cells [6], or a blockade of the androgen negative feedback in hypothalamus and pituitary mechanism [24]. These results show that EDC treatment can lead to hormonal balance disruptions that may or may not be in accordance with the described EDC in vitro mechanism of action. The advantage of measuring the sex steroid concentrations in ovaries is the possibility of evaluating the effects of EDCs on ovarian steroidogenesis. It is important to be aware that in female rats, sex steroids, and almost all estrogens, are produced and secreted by the ovaries. However, there can be external factors influencing hormonal balance; therefore, these gonadal dosages must be followed by a serum measurement of sex steroid concentrations. The obtained ovarian EDC concentrations are of the same order of magnitude as those employed in most in vitro studies of these chemicals (Table 4; [39]). This concentration level provides an important link between our results on ovaries and the results of in vitro studies of corresponding cell types (granulosa and theca cells). Furthermore, because in vitro studies supply more use- ful mechanistic information when they are executed at relevant concentrations and with a good test chemical (parent product or metabolite), this method gives good data for further in vitro study design. However, in vitro studies cannot substitute for in vivo ones, especially because they lack metabolism and other external regulation like the hypothalamo-hypophyseal axis. The need for an integrative evaluation of in vivo and in vitro measurements is reinforced by the observation, in our study, of different in vivo patterns for EDCs with the same described in vitro mechanisms of action. In conclusion, an analytical method was implemented for the determination of sex steroid hormone concentrations and EDC concentrations in rat ovaries. The QuEChERS sample preparation protocol, which is new for this type of matrice, may be qualified as simple (a liquid liquid extraction step followed by a cleaning step), rapid (extractions were performed in less than 15 min) and efficient (RSDs and recoveries were acceptable). The sensitivity and specificity of the LC-MS/MS analysis method allowed the simultaneous detection and quantification of low levels of two classes of molecules: steroids and EDCs. The relevance of this method, validated according to ICH criteria, was confirmed by the application of the methodology to an in vivo study. Thus, as well as providing a new tool in the endocrine toxicology field, this work provided new information about the toxicological patterns of ...
Context 3
... steroid concentrations in the ovaries are presented in Fig. 6. After a 2-week treatment with ATZ, females showed slightly increased ovarian estrone. VCZ treatment significantly increased ovarian androstenedione and estrone. MXC-treated females showed a decrease in androstenedione and testosterone concentrations in the ovaries. BPA treatment showed no significant effect. After the recovery period, there was no more significant effect on sex steroid levels. As far as estrogen-to-androgen ratios are concerned, ATZ and MXC treatments significantly increased the ovarian estrone-to-androstenedione ratio. An over- or under-secretion of one or more steroids can cause a steroidal imbalance that results in various disorders. Precise steroid analysis is very important, not only to predict pathological alterations, but also to understand toxicity mechanisms. Indeed, small variations in steroid concentrations can induce major toxicological changes, particularly when exposure occurs during a critical win- dow [35]. There are many challenges when analyzing steroids, due to their low concentrations and their struc- tural similarities. The method developed herein aimed to simultaneously quantify sex steroids and EDCs in rat ovaries. The excellent recoveries obtained for the sex hormones (Table 2), as compared with the recoveries reported in the literature (higher than 80 %) [10], highlight the efficiency of the “ QuEChERS ” approach for EDCs as well as for hormone extraction. Moreover, the biases of the recoveries as determined by the RSDs are sufficient and acceptable because they are lower than 20 % (Table 2). As ovaries are complex matrices, it was important to thoroughly study the matrix effect aspect. Even if ion suppression was observed, the low variations of recovery, illustrated by low values of matrix effect (Table 2), show the efficiency of the extraction process. To the best of our knowledge, this is the first time that “ QuEChERS ” sample preparation was applied to rat ovaries with analyte concentrations reported in the nanogram range per gram of tissue and the first time that the simultaneous analysis of hormones and EDCs was conducted. LODs and LOQs are similar to those obtained with tissues from other species [10]. Furthermore, the method was sufficiently sensitive and precise to accurately quantify ovarian steroid and EDCs levels in female rats. To better characterize the effects of EDCs in female rats, we focused our work on hormone production in the ovaries after in vivo EDC exposure. We took into account not only the effect on absolute sex steroid concentration, but also on the estrogen-to-androgen ratio and the reproductive side effects. We discuss below the potential mechanisms for altered hormone balance. Following EDC treatment, some females presented estrous cycle disruption and histopatho- logical disorders of the uterus or ovaries (Table 1). These alterations were caused by hormonal balance disruption or altered receptor availability. Ovarian sex steroid concentrations were altered following ATZ, VCZ, and MXC treatments (Fig. 6). ATZ, MXC, and BPA are classified as estrogenic, according to in vitro studies [4, 5, 7]. However, the effects on hormonal balance in our study are more or less important. ATZ elevated the estrone-to-androstenedione ratio by increasing estrone. This effect was in accordance with several other studies, which associated ATZ with disturban- ces in the balance between androgens and estrogens in male rats [36]. This elevated estrogen-to-androgen ratio could be specifically linked to aromatase activity induction observed in in vitro studies [37]. Ovaries from females treated with MXC showed normal estrogen levels and decreased androgen levels. Alteration of steroidogenesis resulting in an androgen decrease has been previously described in male rats following MXC treatment [38]. Thus, MXC appears to exert an anti-androgenic activity in rats. As the effect of BPA on rat ovaries was not significant, we cannot conclude what effect BPA had on the sex steroid levels in our study. VCZ treatment has been associated with anti-androgenic effects in vitro, via the antagonistic binding of its metabolite M2 to the androgen receptor [6]. In our study, VCZ increased ovarian androstenedione and testosterone levels, an effect which is opposite to what is usually reported for this class of compound (anti-androgenic). This effect could account for a direct steroidogenic up-regulation, previously reported in rat and human cells [6], or a blockade of the androgen negative feedback in hypothalamus and pituitary mechanism [24]. These results show that EDC treatment can lead to hormonal balance disruptions that may or may not be in accordance with the described EDC in vitro mechanism of action. The advantage of measuring the sex steroid concentrations in ovaries is the possibility of evaluating the effects of EDCs on ovarian steroidogenesis. It is important to be aware that in female rats, sex steroids, and almost all estrogens, are produced and secreted by the ovaries. However, there can be external factors influencing hormonal balance; therefore, these gonadal dosages must be followed by a serum measurement of sex steroid concentrations. The obtained ovarian EDC concentrations are of the same order of magnitude as those employed in most in vitro studies of these chemicals (Table 4; [39]). This concentration level provides an important link between our results on ovaries and the results of in vitro studies of corresponding cell types (granulosa and theca cells). Furthermore, because in vitro studies supply more use- ful mechanistic information when they are executed at relevant concentrations and with a good test chemical (parent product or metabolite), this method gives good data for further in vitro study design. However, in vitro studies cannot substitute for in vivo ones, especially because they lack metabolism and other external regulation like the hypothalamo-hypophyseal axis. The need for an integrative evaluation of in vivo and in vitro measurements is reinforced by the observation, in our study, of different in vivo patterns for EDCs with the same described in vitro mechanisms of action. In conclusion, an analytical method was implemented for the determination of sex steroid hormone concentrations and EDC concentrations in rat ovaries. The QuEChERS sample preparation protocol, which is new for this type of matrice, may be qualified as simple (a liquid liquid extraction step followed by a cleaning step), rapid (extractions were performed in less than 15 min) and efficient (RSDs and recoveries were acceptable). The sensitivity and specificity of the LC-MS/MS analysis method allowed the simultaneous detection and quantification of low levels of two classes of molecules: steroids and EDCs. The relevance of this method, validated according to ICH criteria, was confirmed by the application of the methodology to an in vivo study. Thus, as well as providing a new tool in the endocrine toxicology field, this work provided new information about the toxicological patterns of ...
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... Eldridge showed in chronic studies, with doses of 25 and 200 mg/kg of atrazine administered orally, they caused an increase in the duration of the estrous cycle and an increase in the number of days in estrus, which was related with the increase in the number of breast tumors in adult female rats [69,70]. Cooper [71] observed that adult rats that Female offspring of rats exposed during pregnancy and/or lactation Higher levels of ADD in plasma, adrenal gland, brain, and breast tissues [74] 200 mg/kg 14 days adult rats Accumulation of atrazine in ovarian tissue [73] 400 mg/kg 14 days adult rats Alteration in the estrous cycle, ovotoxicity and infertility that atrazine can cause infertility, impair folliculogenesis, and alter ovarian function. In addition, an additional toxic mechanism to the increase in progesterone after exposure to atrazine was the reduction in the expression of LH receptors in granulosa cells, which may cause a decrease in LH and estradiol functions in the ovary, including the development of the corpus luteum, the size of ovaries and estrus cyclicity. ...
... A study conducted by Quignot showed that exposing female rats to 200 mg/kg of atrazine for 14 days caused the accumulation of atrazine in ovarian tissue [74]. The results of these studies provide support for the reproductive dysfunction caused by exposure to atrazine and that this substance, as well as its metabolite DDA, is present in brain tissue and breast tissue, supporting the fi ndings that demonstrate that gestational and lactational exposure is harmful to development [8]. ...
... Quignot reported that the eff ects of atrazine on the HHG and HHA axes may culminate in adverse impacts on in vitro, gestational, peripubertal and adult exposures. These changes ranged from puberty delays, altered estrous cycle, atresic ovarian follicles, reduced gonadotropins, and cellular and genetic alterations [74]. These data provide support for considering atrazine as an endocrine disruptor and its ability to cause reproductive dysfunction at various stages of life. ...
Herbicides represent the largest portion of pesticides used both worldwide and in Brazil. Many of these compounds are applied on a large scale in native forests and in urban and industrial water environments, including atrazine. Due to its low cost, ability to remain active in the soil for long periods and potential effect on weed removal, atrazine ranks 5th in the ranking of most used pesticide in Brazil. Although the use of pesticides increases agricultural production, their intensive use can often cause negative effects on fauna and flora. Studies have shown that exposure to atrazine can cause various harmful effects in mammals, of both sexes, such as structural, neuroendocrine and/or behavioral changes. Considering the seriousness of the situation and the possible toxicological and pathological implications that atrazine can generate in the animal organism, the objective of this work was to carry out an integrative literature review in order to verify the scientific panorama on issues related to atrazine exposure and its impacts, mainly with regard to its toxicity on the central nervous system. To carry out this article, a bibliographic survey of scientific material obtained in the following databases was carried out: US National Library of Medicine - National Institutes of Health (PubMed), Virtual Health Library (Latin American and Caribbean Literature in Health Sciences - LILACS), Science Direct and Google® Academic, in the last 25 years. The MeSH Terms used in the search were: “Parkinson's disease”, “atrazine”, “herbicide” and “endocrine disruptor”. The following were found in the Science Direct indexers: 115 records, PubMed 52 records, in LILACS no articles were found, and 1330 records were found in Google® Academic.
... The EDs affect the endocrine system posing a threat to human and animal health and interacting with a receptor of hormones, thus causing several metabolic alterations in the organism (Bila & Dezotti, 2007). Although such contaminants are usually found in the environment at low concentrations, some of them are toxic or cause adverse effects at concentrations of nanograms per liter (Bila & Dezotti, 2007;Newbold et al., 2008;Quignot et al., 2012a;Quignot et al., 2012b;Uchtmann et al., 2020) The drugs are composed of different chemical classes with different toxicity. Antibiotics, in particular, constitute a class of major concern due to their ability to induce and disseminate antimicrobial resistance in the environment. ...
This study aimed to investigate the occurrence of drugs and endocrine disrupters in water supplies and in water for human consumption. Twelve sampling campaigns were carried out during the rainy and dry season at four sampling points in the Bolonha Complex, in the city of Belém, northern region of Brazil: Bolonha reservoir (catchment) and Water Treatment Plant (WTP) Bolonha (filtered water chamber, treated water tank, and washing water from the filters). The determination of the compounds was performed by solid phase extraction followed by gas and liquid chromatography coupled to mass spectrometry. The results confirmed the anthropic influence that the reservoir and WTP-Bolonha have been suffering, as consequence of the discharge of domestic sewage in natura. Among 25 microcontaminants analyzed, 12 were quantified in raw water and 10 in treated water. The antiallergic Loratadine (LRT) was the contaminant that occurred most frequently in all sample points, having been poorly removed (median 12%) in the conventional treatment used. Losartana (LST), 4-octylphenol (4-OP), and Bisphenol A (BPA) also occurred very frequently in raw water with concentrations ranging from 3.7 to 194 ng L⁻¹. Although such contaminants occurred in treated water in concentrations varying from 4.0 to 135 ng L⁻¹, the estimated margin of exposure ranged from 55 to 3333 times which indicates low risk of human exposure to such contaminants through ingestion of treated water.
... Androgen levels were unchanged due to atrazine treatment. In the few published studies that used LC-MS/ MS, there was no evidence of changes in plasma estrogen or androgen concentrations after atrazine treatment ( Quignot et al., 2012b;Tournier et al., 2015). Quignot et al. (2012b) extracted samples directly from ovaries of atrazine-treated rats (200 mg/kg for 14 days) and found no change in androstenedione, testosterone, or estradiol, with a small rise in estrone ( Quignot et al., 2012b). ...
... In the few published studies that used LC-MS/ MS, there was no evidence of changes in plasma estrogen or androgen concentrations after atrazine treatment ( Quignot et al., 2012b;Tournier et al., 2015). Quignot et al. (2012b) extracted samples directly from ovaries of atrazine-treated rats (200 mg/kg for 14 days) and found no change in androstenedione, testosterone, or estradiol, with a small rise in estrone ( Quignot et al., 2012b). Tournier et al. (2015) reported no effect of atrazine on absolute sex hormone levels, but found an elevation of estradiol/testosterone ratio in females. ...
... In the few published studies that used LC-MS/ MS, there was no evidence of changes in plasma estrogen or androgen concentrations after atrazine treatment ( Quignot et al., 2012b;Tournier et al., 2015). Quignot et al. (2012b) extracted samples directly from ovaries of atrazine-treated rats (200 mg/kg for 14 days) and found no change in androstenedione, testosterone, or estradiol, with a small rise in estrone ( Quignot et al., 2012b). Tournier et al. (2015) reported no effect of atrazine on absolute sex hormone levels, but found an elevation of estradiol/testosterone ratio in females. ...
T cell-dependent IgM antibody production and natural killer cell (NKC) activity were assessed in SD rats orally administered atrazine for 28 days to males (0, 6.5, 25, or 100 mg/kg/day) or females (0, 3, 6, or 50 mg/kg/day), or 30 or 500 ppm in diet (3 or 51 mg/kg/day). Anti-asialo GM1 antibodies (NKC) and cyclophosphamide (antibody-forming cell [AFC]) assays served as positive controls. Pituitary (ACTH, prolactin), adrenal (corticosterone, progesterone, aldosterone), and gonadal (androgens, estrogens) hormones were assessed after 1, 7, and/or 28 days of treatment. Food intake and body weights were significantly reduced in the highest dosed males, and transiently affected in females. Urinary corticosterone levels were not increased in atrazine-treated groups in either sex at any time point measured (10, 22, or 24 days). Corticosterone and progesterone were elevated in males after a single atrazine dose ≥6.5 mg/kg/day, but not after 7, 14, or 28 doses. There were no effects on adrenal, pituitary, or gonadal hormones in females. Atrazine did not suppress the AFC response or decrease NKC function after 28 days in males or females. Atrazine had no effect on spleen weights or spleen cell numbers in males or females, although thymus weights were elevated in males receiving the highest dose. The lack of immunotoxic effect of atrazine was associated with diminished adrenal activation over time in males, and no effects on adrenal hormones in females.
... pressurized liquid extraction (Du et al. 2012;Zhao et al. 2015a) and fluorimetry (Pacioni and Veglia 2003). For pesticide extraction procedure, several methodologies are used, such as QuEChERS (Quignot et al. 2012;Martínez-Domínguez et al. 2015;Paz et al. 2015) and liquid-solid extraction (LSE) (Wang et al. 2011b;Łozowicka et al. 2014), which have been reviewed comprehensively in the literature (García-Valcárcel and Tadeo 2009;Kolberg et al. 2011). To detect pesticide residues, GC and LC are frequently employed (Páleníková et al. 2015; Barceló et al. 1995). ...
This review discusses the past and recent advancements of biosensors focusing on detection of organophosphorus pesticides (OPs) due to their exceptional use during the last decades. Apart from agricultural benefits, OPs also impose adverse toxicological effects on animal and human population. Conventional approaches such as chromatographic techniques used for pesticide detection are associated with several limitations. A biosensor technology is unique due to the detection sensitivity, selectivity, remarkable performance capabilities, simplicity and on-site operation, fabrication and incorporation with nanomaterials. This study also provided specifications of the most OPs biosensors reported until today based on their transducer system. In addition, we highlighted the application of advanced complementary materials and analysis techniques in OPs detection systems. The availability of these new materials associated with new sensing techniques has led to introduction of easy-to-use analytical tools of high sensitivity and specificity in the design and construction of OPs biosensors. In this review, we elaborated the achievements in sensing systems concerning innovative nanomaterials and analytical techniques with emphasis on OPs.
... ATR has been detected in human follicular fluid at a very low, residual concentration [46]. Quignot and collaborators have detected 0.5 M ATR in rat ovary following oral administration of 200 mg/kg bw per day for two weeks [47]. Since the authors did not measure the ATR metabolites, it remains unknown whether ATR was metabolized in the ovary or whether the ovarian effects of ATR came from the parent compound or its metabolites. ...
... These lower levels allow for a more translatable approach to epidemiological studies (discussed in Section 5). A second study conducted by Quignot et al., exposed female Sprague-Dawley rats to 200 mg/kg atrazine for 14 days and found an accumulation of atrazine in ovarian tissue [73]. Results from these studies provide support for the reproductive dysfunction elicited by atrazine exposure through multiple avenues. ...
... In addition, the identification of atrazine in the adrenals provides support that atrazine can also target additional neuroendocrine pathways including the HPA axis. Although gonadal tissue requires intensive sample preparation due to their complex biological matrix and high lipid content, these studies show that atrazine is present in ovarian tissue [73] and highlight that further work is needed to link ovarian atrazine levels to observed histological, hormonal, and cellular alterations. ...
Endocrine disrupting chemicals (EDC) are exogenous agents that alter endogenous hormone signaling pathways. These chemicals target the neuroendocrine system which is composed of organs throughout the body that work alongside the central nervous system to regulate biological processes. Of primary importance is the hypothalamic-pituitary-gonadal (HPG) axis which is vital for maintaining proper reproductive function. Atrazine (2-chloro-4-ethylamino-6-isopropylamino-1,3,5-triazine) is a pre-emergent herbicide used to prevent the growth of weeds on various crops. This herbicide is reported to widely contaminate potable water supplies everywhere it is applied. As such, the European Union banned the use of atrazine in 2004. Currently the United States Environmental Protection Agency regulates atrazine at 3 parts per billion (ppb; μg/L) in drinking water, while the World Health Organization recently changed their drinking water guideline to 100 ppb. Atrazine is implicated to be an EDC that alters reproductive dysfunction by targeting the HPG axis. However, questions remain as to the human health risks associated with atrazine exposure with studies reporting mixed results on the ability of atrazine to alter the HPG axis. In this review, the current findings for atrazine’s effects on the HPG axis are examined in mammalian, anuran, and fish models and in epidemiological studies.
... This method is based on extraction in acetonitrile followed by dispersive solid-phase extraction (DSPE), the entire method being performed in two centrifugation tubes. QuEChERS quantification methods are commonly applied to food, water and grass but this approach is only rarely applied to biological samples [23][24][25][26]. ...
... This method has also been applied to gram-scale tissue samples, for the determination of four endocrine disruptors (bisphenol A, atrazine, and the active metabolites of methoxychlor and vinclozolin) in rat ovaries and testis [25,26]. ...
... LC-MS is more commonly applied for the analysis of hormones. There are a number of papers on hormone determination in biological [49][50][51][52][53][54][55], and environmental matrices, such as water [26,56,57] and sediments [9,[58][59][60][61][62][63][64][65]. Electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) are both equally employed for ionization of hormones and sterols. ...
In this paper, development and optimization of new LC-MS method for determination of twenty selected hormones, human/animal and plant sterols in river sediments were described. Sediment samples were prepared using ultrasonic extraction and clean up with silica gel/anhydrous sodium sulphate cartridge. Extracts were analyzed by liquid chromatography-linear ion trap-tandem mass spectrometry, with atmospheric pressure chemical ionization. The optimized extraction parameters were extraction solvent (methanol), weight of the sediment (2g) and time of ultrasonic extraction (3× 10min). Successful chromatographic separation of hormones (estriol and estrone, 17α- and 17β-estradiol) and four human/animal sterols (epicoprostanol, coprostanol, α-cholestanol and β-cholestanol) that have identical fragmentation reactions was achieved. The developed and optimized method provided high recoveries (73-118%), low limits of detection (0.8-18ngg(-1)) and quantification (2.5-60ngg(-1)) with the RSDs generally lower than 20%. Applicability of the developed method was confirmed by analysis of six river sediment samples. A widespread occurrence of human/animal and plant sterols was found. The only detected hormone was mestranol in just one sediment sample.
... So it is important to develop analytical methods to measure free concentrations in the medium, cellular concentrations and quantity bound to the vial walls or medium proteins over time. Automation and analytical methods such as liquid chromatography with tandem mass spectroscopy (LC/MS/MS) have vastly increased the sample throughput for in vitro ADME assays [ 65 ], and the same should hold for in vitro toxicity assessment [ 66 ]. Metabolomic data might be an answer to that question but keeping in mind that enough time points have to be available to reconstruct concentration vs. time curves. ...
The increasing use of in vitro systems in pharmacology and toxicology has the potential to yield high-throughput screening of molecules and in-depth mechanistic evaluations of toxicity. However, the relevance of results obtained from simplified systems to humans may be questionable. To address that issue, consistent and reliable extrapolation procedures from in vitro results to human in vivo are needed. Developing those procedures requires to first understand the basis and limitations of in vitro experiments as well as the needs of risk assessment and safety evaluation. This chapter gives an overview of the strategies used for quantitative in vitro to in vivo extrapolations in pharmacokinetics (PK), pharmacodynamics (PD), up to the PK/PD continuum. Their scientific and technical challenges are also discussed.
... Therefore, diminished level of LHR in atrazineexposed granulosa cells appears to lead to general reduction in the signaling via EGFR and Pgr networks. Recent in vivo study by Quignot et al. (2012b) shows that 14-day exposure of adult rats to atrazine led to accumulation of the parent compound but not atrazine metabolites in the ovarian tissue. There, atrazine decreases LHR-and estradioldependent functions in the ovary such as maintenance and development of the corpora lutea, the size of the ovaries and the normal estrus cycles (Quignot et al., 2012b). ...
... Recent in vivo study by Quignot et al. (2012b) shows that 14-day exposure of adult rats to atrazine led to accumulation of the parent compound but not atrazine metabolites in the ovarian tissue. There, atrazine decreases LHR-and estradioldependent functions in the ovary such as maintenance and development of the corpora lutea, the size of the ovaries and the normal estrus cycles (Quignot et al., 2012b). These data provide an important link between the in vivo effect of atrazine on the adult ovaries and the mechanisms of atrazine action obtained from this and other in vitro studies. ...
... These data provide an important link between the in vivo effect of atrazine on the adult ovaries and the mechanisms of atrazine action obtained from this and other in vitro studies. Therefore, adult exposure to atrazine leads to accumulation of the parent compound in the ovarian cells where it suppresses the FSH-induced LHR transcription (this study) and changes the balance of the steroid hormones (Quignot et al., 2012a(Quignot et al., , 2012b, thus negatively affecting ovulation and female fertility. ...
