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International audience; With the objectives of both generating bisphenols (BPs) conjugates occurrence data in food from animal origin but also investigating the origin of associated contamination, the present study deals with the development of an efficient analytical method aiming at monitoring both BPA and BPS conjugated metabolites in food from animal origin. The objective of such monitoring is to determine the origin of BPs contamination (FCM or animal contamination). The targeted compounds were BPA-monoglucuronide (BPA-1G), BPA-diglucuronide (BPA-2G), BPA-monosulfate (BPA-1S), BPA-disulfate (BPA-2S) and BPS-monoglucuronide (BPS-1G). The developed standard operating procedure includes a preliminary solid-liquid extraction step followed by two successive solid phase extraction (SPE) stages, using successively a non-polar phase and a strong cation exchange polymer. Quantification was achieved according to both the isotopic dilution and surrogated quantification methods, using 13C-BPA-1G and BPA-d6-1S as internal standards. Linearity was validated (R2 > 0.99) for each molecule within the concentration range [0-10] μg kg-1. Detection limits ranged from 0.02 μg kg-1 (BPA-1G in muscle, BPA-1S and BPA-2G in liver) to 0.50 μg kg-1 (BPA-2S in muscle). The strategy was then proven on liver samples collected from pregnant ewes subcutaneously exposed to BPA during 105 days, at 50 μg kg-1 per day. BPA-1G, BPA-2G and BPA-1S were detected and quantified at a concentration of 3.81 μg kg-1, 0.80 μg kg-1 and 0.09 μg kg-1, respectively. The analytical method was finally implemented on fifty unpacked food samples from animal origin in which significant free BPA concentrations were previously measured. Since no metabolites of BPA could be measured (

Bisphenol S (BPS) is widely used as a substitute for Bisphenol A in consumer products. Despite its potential endocrine-disrupting effects and widespread exposure, toxicokinetic data, particularly during the critical period of pregnancy, are not available for BPS. The objectives of our study were to evaluate the mechanisms determining fetal exposure to BPS and to BPS glucuronide (BPSG) and to compare them with those prevailing for BPA.The disposition of BPS and BPSG was evaluated in the materno-fetal unit of the catheterized pregnant ewe model, following intravenous administrations of BPS and BPSG to mothers and their fetuses. In a second experiment, the rate of BPS accumulation in the fetal compartment was determined under steady-state conditions after repeated intravenous BPS administrations to the mother.In the maternal compartment, BPS was mainly metabolized into BPSG and totally eliminated in urine. Only 0.40% of the maternal dose was transferred to the fetus. However, once in the fetal compartment, 26% of the fetal dose was rapidly eliminated through placental transfer, while 46% of BPS was metabolized into BPSG which remained trapped in the fetal compartment. Thus, the elimination of BPSG from the fetal compartment required its back-conversion into bioactive BPS, leading to an 87% enhancement of the fetal BPS exposure.Our findings demonstrate that, despite the low materno-fetal placental transfer of BPS, this substitute for BPA is able to accumulate in the fetal compartment after repeated maternal exposure, leading to chronic fetal exposure to BPS in a range of concentrations similar to those of BPA. Keywords: Bisphenol S, Bisphenol S Glucuronide, Toxicokinetic, Fetal exposure, Bisphenol A analogue

Background: Given its hormonal activity, bisphenol S (BPS) as a substitute for bisphenol A (BPA) could actually increase the risk of endocrine disruption if its toxicokinetic (TK) properties, namely its oral availability and systemic persistency, were higher than those of BPA. Objectives: The TK behavior of BPA and BPS was investigated by administering the two compounds by intravenous and oral routes in piglet, a known valid model for investigating oral TK. Methods: Experiments were conducted in piglets to evaluate the kinetics of BPA, BPS, and their glucuronoconjugated metabolites in plasma and urine after intravenous administration of BPA, BPS, and BPS glucuronide (BPSG) and gavage administration of BPA and BPS. A population semiphysiologically based TK model describing the disposition of BPA and BPS and their glucuronides was built from these data to estimate the key TK parameters that drive the internal exposure to active compounds. Results: The data indicated that almost all the BPS oral dose was absorbed and transported into the liver where only 41% of BPS was glucuronidated, leading to a systemic bioavailability of 57.4%. In contrast, only 77% of the oral dose of BPA was absorbed and underwent an extensive first-pass glucuronidation either in the gut (44%) or in the liver (53%), thus accounting for the low systemic bioavailability of BPA (0.50%). Due to the higher systemic availability of BPS, in comparison with BPA, and its lower plasma clearance (3.5 times lower), the oral BPS systemic exposure was on average about 250 times higher than for BPA for an equal oral molar dose of the two compounds. Conclusion: Given the similar digestive tracts of pigs and humans, our results suggest that replacing BPA with BPS will likely lead to increased internal exposure to an endocrine-active compound that would be of concern for human health. https://doi.org/10.1289/EHP4599

Refers to: Jeremy Gingrich, Yong Pu, Richard Ehrhardt, Rajendiran Karthikraj, Kurunthachalam Kannan, Almudena Veiga-Lopez Toxicokinetics of bisphenol A, bisphenol S, and bisphenol F in a pregnancy sheep model Chemosphere, Volume 220, April 2019, Pages 185-194; International audience; Gingrich et al. (2019) report a comparative toxicokinetic (TK) study of Bisphenol A (BPA), Bisphenol S (BPS) and Bisphenol F (BPF) in the pregnant ewe model after a single subcutaneous administration of BPS or a mixture of BPA, BPS and BPF. The topic is highly relevant. Unfortunately, many errors were noted in the calculation of TK parameters. In the results reported, the disposition of aglycone is systematically confused with that of the corresponding glucuronide, and the interpretation of computed or observed data is therefore erroneous. This makes the paper misleading in a field that does not need these kinds of error to fuel debates that are already heated and confusing. Inspection of Table 2 immediately indicates that the computed clearances (CL) of 45.4 mL/h for BPA (but actually BPA plus BPAG) or the clearance of 5.2 mL/h for BPS (but actually BPS and BPSG) are impossible from a biological point of view. Indeed, this is clearly illustrated by the estimation of Volume of Distribution (Vss) from the product of Mean Residence Time (MRT) and Clearance (CL) (Toutain and Bousquet-Melou, 2004). The computed MRTs of 8.3 h for BPA and 6.7 h for BPS produce an estimated Vss of 4.94 and 0.457 mL/kg for BPA and BPS, respectively. However, these values are not plausible, given that the minimal possible Vss for any substance is the plasma volume (i.e. about 40 mL/kg in sheep). This error in the computation of plasma clearance is rooted in the flawed computation of the corresponding AUC (0.81 and 7.5 mg*h/mL for BPA and BPS, respectively). The order of magnitude of these AUCs can be readily obtained by dividing the maximal observed plasma concentration (Cmax of 66.7 and 643 ng/mL for BPA and BPS, respectively) by the slope of the terminal phase (that is equal to 0.693/half-life) (Toutain and Bousquet-Mélou, 2004). This leads to computed AUCs of 0.000510 and 0.00343 mg*h/mL for BPA and BPS, respectively. These roughly estimated AUCs are 1588–2185 fold lower than those reported in the present article and we suspect that the authors, at the very least, made an error in handling their concentration units. Using correctly computed AUCs would lead to estimates of total BPA and BPS plasma clearance (actually CL/F, with F the bioavailability factor associated with the subcutaneous administration) of about 980 and 146 mL/kg/h. These values would be of the same order of magnitude as the plasma clearance that we directly measured using an IV administration of BPAG and BPSG in sheep (333 and 49 mL/kg/h) for BPAG and BPSG (Gauderat et al., 2016, Grandin et al., 2018). The remaining difference is probably due in part to incomplete bioavailability following the subcutaneous administration of BPA and BPS. Moreover, the authors wrote in their “Toxicokinetic analysis” section that the TK parameters were calculated using the plasma concentrations of total BPA, BPS, or BPF (i.e. unconjugated and conjugated forms of bisphenols). Bisphenol metabolites and bisphenol aglycone are distinct substances with their own physico-chemical properties and their own dispositions. Therefore, total bisphenol concentrations cannot be considered for a meaningful TK analysis. Given that the ratio of the bisphenol metabolite over bisphenol aglycone is about 100 for BPA (Gauderat et al., 2016) and 10 for BPS (Grandin et al., 2018), the results published by Gingrich et al. describe the disposition of the bisphenol metabolites i.e. BPAG and BPSG rather than the disposition of bisphenol aglycone, at least for BPA and BPS. Hence the main message conveyed by the present paper is not related to BPA and BPS but to their mainly inactive metabolite. Other surprising results are the urinary concentrations. It is claimed that 9 h after bisphenol administration, the peak maternal urine concentrations for BPA and BPS were 1300 and 3870 ng/mL, respectively. Again, this is very unlikely as the daily urine output in sheep is similar to that of humans (i.e. 1.5 L for sheep of 76.3 Kg BW). If, as a worst-case scenario, we consider that these so-called maximal urine concentrations are similar to the average urine concentration, the total amounts of BPA/BPAG and BPS/BPSG that are eliminated by urine over 24 h are no more than 2 and 6 mg in total, i.e. about 5 and 16% of the administered dose of 36.7 mg, for BPA/BPAG and BPS/BPSG, respectively. This is at variance with the literature reports that most of the BPA/BPAG is eliminated in urine over 24 h (Gauderat et al., 2016). The discrepancy of the authors’ result may be due to the low bioavailability of the two tested compounds or, more likely, to the fact that most of the dose had already been eliminated during the first hours following subcutaneous administration and was simply missed by the sampling design because the first urine sampling at 9 h post-administration was too late. The interpretation of the results is also questionable. For instance, the authors suggest that a first-pass effect may reduce the terminal half-life of a substance or compare MRTs (as computed for an extravascular route with a mixture of the aglycone and its conjugate) with a half-life, as obtained after an IV administration. The authors may not have realized that MRT are additives, not half-lives, and that what they actually measured were not the genuine MRTs of the different substances under investigation (only possible by an IV administration) but an overall MRT that includes a Mean Absorption time (MAT) plus a Mean Time of conjugation of bisphenol aglycone into the bisphenol metabolite. This misunderstanding makes any comparison with the results obtained for Bisphenols by IV route meaningless. Finally, the authors indicate in their conclusion that “this is the first report on the toxicokinetic of either BPS and BPF in a pregnancy model”. This statement is, to our knowledge, correct for BPF but, for BPS, the first toxicokinetic parameters were published in Environment International and have been available since August 2018 (Grandin et al., 2018).

International audience; Previous data obtained in piglets suggested that despite structural analogy with Bisphenol A (BPA), Bisphenol S (BPS) elimination may proceed more slowly, resulting in a much higher systemic exposure to unconjugated BPS than to BPA. Interspecies allometric scaling was applied to predict the toxicokinetic (TK) parameters of BPS, namely plasma clearance in humans from values obtained in animals, and thus contribute to assessment of the human internal exposure to BPS. Allometric scaling was performed using mean BPS plasma clearance values measured in rats after intravenous administration of 5 mg BPS /kg body weight (BW) and those previously obtained in piglets and sheep using identical IV BPS dosing and analytical procedures. The BPS plasma clearance, evaluated at 0.92 L/kg.h in rats, was proportional to species body weight, enabling the prediction of human BPS plasma clearance by extrapolating to a BW of 70 kg. The estimated BPS plasma clearance in humans was thus 0.92 L/min (0.79 L/kg.h), i.e. about two times lower than the previously estimated BPA clearance (1.79 L/min). By increasing systemic exposure to the active moiety of an environmental estrogenic chemical, this less efficient clearance of BPS in humans, as compared with BPA, might worsen the harmful consequences of replacing BPA by BPS.

International audience; The aim of our study was to evaluate the bidirectional transfer of Bisphenol S (BPS) and its main metabolite, BPS Glucuronide (BPSG), using the model of perfused human placenta and to compare the obtained values with those of Bisphenol A (BPA) and BPA Glucuronide. Fourteen placentas at term were perfused in an open dual circuit with deuterated BPS (1 and 5 μM) and non-labelled BPSG (2.5 μM) and a freely diffusing marker antipyrine (800 ng/ml) in the presence of albumin (25 mg/ml). In a second experiment, the potential role of P-glycoprotein in the active efflux of BPS across the placental barrier was studied using the well-established P-glycoprotein inhibitor, PSC833 (2 and 4 μM). Placental transfer of BPS was much lower than that of BPA in both directions. The placental clearance index of BPS in the materno-fetal direction was three times lower than in the opposite direction, strongly suggesting some active efflux transport. However, our results show that P-glycoprotein is not involved in limiting the materno-fetal transfer of BPS. Placental transfer of BPSG in the fetal compartment was almost non-existent indicating that, in the fetal compartment, BPSG originates mainly from feto-placental metabolism. The feto-maternal clearance index for BPSG was 20-fold higher than the materno-fetal index. We conclude that the blood-placental barrier is much more efficient in limiting fetal exposure to BPS than to BPA, indicating that the placenta has a crucial role in protecting the human fetus from BPS exposure.

International audience

International audience; Regulatory measures and public concerns regarding bisphenol A (BPA) have led to its replacement by structural analogs, such as Bisphenol S (BPS), in consumer products. At present, no toxicokinetic investigations have been conducted to assess the factors determining human internal exposure to BPS for subsequent risk assessment. Toxicokinetic studies require reliable analytical methods to measure the plasma concentrations of BPS and its main conjugated metabolite, BPS - glucuronide (BPS - G). An efficient on - li ne SPE - UPLC - MS/MS method for the simultaneous quantification of BPS and BPS - G in ovine plasma was therefore developed and validated in accordance with the European Medicine s Agency guidelines for bioanalytical method validation. This method has a limit of quantification of 3 ng mL - 1 for BPS and 10 ng mL - 1 for BPS - G , a n analytical capacity of 200 samples per day, and is particularly well suited to toxicokinetic studies. Use of this method in toxicokinetic studies in sheep showed that BPS, like BPA, is efficiently metabolized into its glucuronide form . However, the clearance s and distribution s of BPS and BPS - G were lower than those of the corresponding unconjugated and glucuroconjugated forms of BPA .