# Prenatal exposures to persistent organic pollutants and menstrual characteristics in girls at age 12 in the French PELAGIE cohort

Maximilien Génard-Walton, Elisabeth Angot, Charline Warembourg, Christine Monfort, Florence Rouget, Nathalie Costet, Hélène Tillault, Fabrice Lainé, Éric Gaudreau, Sylvaine Cordier, et al.

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Maximilien Génard-Walton, Elisabeth Angot, Charline Warembourg, Christine Monfort, Florence Rouget, et al. Prenatal exposures to persistent organic pollutants and menstrual characteristics in girls at age 12 in the French PELAGIE cohort. Environmental Research, 2025, 285 (Part 3), pp.122445. 10.1016/j.envres.2025.122445. hal-05203599

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# Prenatal exposures to persistent organic pollutants and menstrual characteristics in girls at age 12 in the French PELAGIE cohort

Maximilien Génard-Walton<sup>a, *, 1 ID</sup>, Elisabeth Angot<sup>a, 1</sup>, Charline Warembourg<sup>a ID</sup>, Christine Monfort<sup>a ID</sup>, Florence Rouget<sup>b</sup>, Nathalie Costet<sup>a ID</sup>, Hélène Tillault<sup>a</sup>, Fabrice Lainé<sup>c</sup>, Éric Gaudreau<sup>d ID</sup>, Sylvaine Cordier<sup>a</sup>, Marina Kvaskoff<sup>e ID</sup>, Cécile Chevrier<sup>a, 2 ID</sup>, Ronan Garlantézec<sup>b, 2 ID</sup>

## ARTICLE INFO

### Keywords:

[max 7]: persistent organic pollutants
Adolescence
Menarche
Menstruation Disturbances
Dysmenorrhea
Chemical mixtures

## ABSTRACT

Few studies have evaluated the impact of prenatal exposure to persistent organic pollutants (POPs) on menstruation characteristics. Therefore, the aim of our study was to investigate the associations between cord blood concentrations of POPs and menstruation characteristics including menarche status, menstrual regularity, pain and duration in girls at age 12.

We used data from the French PELAGIE mother-child cohort follow-up at 12 years old which included a questionnaire on menstruation characteristics. POP concentrations of 14 organochlorine pesticides (OCPs), 17 polychlorinated biphenyls (PCBs), 5 polybrominated diphenyl ethers (PBDEs), and 9 per-polyfluoroalkyl substances (PFAS) were measured on cord blood sampled at birth. We used multivariate logistic regression and Poisson regression to study the relation between prenatal POPs and menarche status, regularity of menstruations, menstruation duration, and severe pain in the lower abdomen or cramps (N = 296). The effect of mixtures of POPs on these outcomes were assessed using quantile g-computation.

The majority of POPs were associated with a non-statistically significant increased risk of menarche at 12 years old, especially for the PCB mixture (OR = 1.29, 95 % CI [0.99; 1.68]). We also observed a statistically significant association between the second tercile of dieldrin (OR = 2.91, 95 % CI [1.13; 7.82]) and PFNA (OR = 3.13, 95 % CI [1.01; 10.43]) and severe pain in the lower abdomen or cramps, and a statistically significant association between the second tercile of hexachlorobenzene and menstruation duration (% change = 25.6 %, 95 % CI [1.2; 56.6]). For others POPs and other menstrual characteristics we did not observe any associations.

Further longitudinal studies are needed to confirm these results, possibly through combined analyses of several mother-child cohorts to increase study power.

## 1. Introduction

The prenatal period is particularly vulnerable to stressors that may induce structural or functional anomalies visible at birth or later in life

(Bianco-Miotto et al., 2017). The development of the reproductive system starts early during pregnancy and is largely guided by hormones, and may be vulnerable to exogenous chemicals such as endocrine disruptors (EDCs) (Ho et al., 2017). Among EDCs, persistent organic

<footnote><sup>a</sup> Univ Rennes, Inserm, EHESP, Irset (Institut de recherche en santé, environnement et travail), UMR_S 1085, F-35000, Rennes, France</footnote>

<footnote><sup>b</sup> Univ Rennes, CHU Rennes, Inserm, EHESP, Irset (Institut de recherche en santé, environnement et travail), UMR_S 1085, F-35000, Rennes, France</footnote>

<footnote><sup>c</sup> CHU Rennes, Inserm, CIC1414, Rennes, France</footnote>

<footnote><sup>d</sup> Centre de Toxicologie du Québec (CTQ), Institut national de santé publique du Québec (INSPQ), Québec, Canada</footnote>

<footnote><sup>e</sup> Université Paris-Saclay, UVSQ, Inserm, Gustave Roussy, Équipe "Exposome Et Hérédité", CESP UMR1018, 94805, Villejuif, France</footnote>

<footnote>* Given their role as Editor, Charline Warembourg had no involvement in the peer review of this article and had no access to information regarding its peer review. Full responsibility for the editorial process for this article was delegated to another journal editor.</footnote>

<footnote>* Corresponding author. 9 Avenue du Professeur Léon Bernard, 35000, Rennes, France.</footnote>

<footnote>E-mail address: maximilien.genard-walton@inserm.fr (M. Génard-Walton).</footnote>

<footnote n="1">¹ Co-first authors.</footnote>

<footnote n="2">² Co-last authors.</footnote>
pollutants (POPs) are of particular interest as they have been shown to impact the development of the reproductive system (Gore et al., 2015). POPs are chemical substances comprising carbon atoms and halogenated compounds like chlorine, bromine or fluorine atoms, and are grouped into different chemical families including organochlorine pesticides (OCPs), polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs) and per-polyfluoroalkyl substances (PFAS). Although their use in industrial processes have been restricted by national and international institutions (Stockholm convention, 2004), a large proportion are still largely detected in populations across the world due to their persistence in the environment, capacity to travel long distances, and lipophilic properties (except for PFAS which are lipophobic and amphiphilic). Moreover, some of these molecules (PFAS) are still being used in the manufacturing of consumer products including non-stick cookware, grease-resistant food packaging, leather products, and textiles that enhance water, grease, and soil repellence. (Guo et al., 2019; Hsu et al., 2022).

The onset of menstruation (menarche) is a milestone in the girl pubertal development which usually occurs between the ages of 12 and 13 (American Academy of Pediatrics and Committee on Adolescence, 2006; INSERM, 2007). Menstrual cycles typically last 21-34 days in adult women but this interval can be larger in adolescent girls. In adolescents, menstruation duration is usually between one and seven days (Harley et al., 2024). Earlier age at menarche has been associated with various adverse health outcomes such as adult obesity, diabetes, breast cancer, and worse cardiovascular health (Chen et al., 2024) whilst irregular and long menstrual cycles have been associated with greater risks of ovarian cancer, coronary heart disease, type 2 diabetes, and mental health problems (Wang et al., 2020). Moreover, studies have reported associations between early menarche, menstrual cycle length, and chronic pelvic pain, and an increased risk of endometriosis in adolescents (Janssen et al., 2013; Shafrir et al., 2018).

To our knowledge, little research has focused on associations between prenatal exposure to POPs and menstrual cycle characteristics. Among studies investigating the effects of prenatal POP exposure on the age of menarche, PCBs and p,p'-DDE has been associated with a decreased age at menarche (Marks et al., 2021; Vasiliu et al., 2004), and PBDE and PFAS exposure have been associated with a later age at menarche (Harley et al., 2017; Kristensen et al., 2013; Liu et al., 2023). Comparing girls from the Yucheng cohort aged 16-17 born to mothers exposed to the 1978-79 mass PCBs-polychlorinated dibenzofurans (PCDFs) poisoning from cooking oil in central Taiwan (N = 17) and unexposed girls (N = 16), Yang et al. (2005) found shorter bleeding duration assessed by a self-reported diary during 84 days in exposed girls (unadjusted association). In a Danish mother-child cohort established in 1988-89, Kristensen et al. found that prenatal perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS) were not associated with cycle length in girls at age 16 (2013), and prenatal dichlorodiphenyldichloroethylene (p,p'-DDE), hexachlorobenzene (HCB) and a sum of PCBs, were not associated with cycle length in women at age 20 (2016). Menstrual pain and regularity were not assessed in this cohort.

Considering the scarcity on this topic, our objective was to investigate the associations between cord blood concentrations of POPs and menarche status as well as menstruation characteristics including regularity, pain and duration in girls at age 12.

# 2. Materials and methods

## 2.1. Study population

PELAGIE is a mother-child cohort that included 3421 women from the general population from three counties of Brittany, western France, at the beginning of their pregnancy (<19 weeks of gestation) between 2002 and 2006 (Warembourg et al., 2024). Women were recruited by gynaecologists, obstetricians, and hospital and private sonographers and

were followed-up until the end of pregnancy. Women filled out an inclusion questionnaire including information on sociodemographic characteristics, diet, life habits, alcohol consumption and smoking during pregnancy. At birth, blood cord samples were collected for approximately 65 % of the cohort. After centrifugation, serums were frozen at -20 °C in 2 mL tubes. When the children born within this cohort reached 12 years old, a health and environment questionnaire were sent to eligible families which included information on the child's health (diagnoses pathologies, allergies, respiratory health), the condition of their homes, and their dietary habits. Concomitantly, a questionnaire on "Growing and Changing" covering questions on height, weight, menstrual characteristics, pubertal development, and daily habits (physical activity, sleep, and energy levels) were sent to the families to be filled out by the child. Among 2520 eligible families, 1197 complete child questionnaires were returned, including 598 from girls (median [inter-quartile range], minimum, maximum age of response: 12.8 [12.7-12.9], 12.0, 13.0, years old).

## 2.2. Menstrual cycle characteristics

Girls were asked in the "Growing and Changing" questionnaire: "have you ever had your period?" (possible answers: "Yes", "No"). For girls who had started menstruating, girls were asked about their age, month and year of first menstruation, the regularity of their periods (possible answers: "Yes", "No"), menstrual cycle length (in days, only for girls with regular menstruation), menstruation duration (in days), the presence of severe and prolonged bleeding (possible answers: "No", "Yes and I saw a doctor", "Yes and I didn't see a doctor") and the occurrence of severe menstrual pain in the lower abdomen or cramps (possible answers: "No", "Yes and I saw a doctor", "Yes and I didn't see a doctor"). For the final two variables, the condition was considered present if they answered "Yes and I saw a doctor" or "Yes and I didn't see a doctor".

## 2.3. Analyses of POP on cord blood serum samples

Among the 598 girls included in this follow up, measures of prenatal POPs were available for 296 except for PFAS which were measured at a later stage for 218 girls (on the remaining serum samples available). All POP determinations were carried out by the Center of Toxicology of Quebec (CTQ) of the National Institute of Public Health (INSPQ), with gas chromatography-mass spectrometry (GC-MS) for PCBs, OCPs and PBDEs and with liquid chromatography-tandem mass spectrometry (LC-MS/MS) for PFAS. Two mL of cord serum samples were enriched with internal labeled standards. POP compounds (PCBs, OCPs and PBDEs) were extracted by a solid-phase extraction (before 2013) or by a liquid-liquid extraction in the presence of a saturated ammonium sulfate solution (from 2013). The change in extraction methods showed no impact on the quality of the results provided or on the distributions of measurements, as the methods were comparable. These extracts were cleaned up on deactivated 0.5 % florisil columns, evaporated to dryness and the PCBs and OCPs (taken up in hexane) were then analysed on a GC-MS (Agilent Technologies Inc.; Mississauga, Ontario, Canada) with an Agilent 60 m DB-XLB column (0.25 mm i.d., 0.25 µm film thickness), whereas PBDEs were analysed on a GC-MS with a 15 m DB-XLB column (0.25 mm i.d., 0.1 µm film thickness). For both analytic instruments, the carrier gas was helium and all injections were in splitless mode. The MS used single ion monitoring (SIM) and negative chemical ionization (NCI) with methane as the reagent gas. The PFAS analysis was performed as follows: 100 µL of cord serum samples were enriched with ¹³C-labeled internal standards and were acidified with a 50 % formic acid solution. Thereafter, the samples were extracted using a solid phase extraction (SPE) with Silia PrepX WAX cartridges 100 mg/3 mL (Silicycle; Québec, Canada). The extracts were evaporated to dryness and dissolved in 1 mL of 5 mM ammonium in 20 % acetonitrile before to be analysed by Ultra Performance Liquid Chromatography (UPLC Waters Acquity) with a tandem mass spectrometer (MS/MS Waters Xevo TQ-S) (Waters;
Milford, MA, USA) in the MRM mode with an electrospray ion source in the negative mode. The column used was an ACE EXCEL C₁₈-PFP 100 mm × 2.1 mm, 2.0 µm (ACE; Aberdeen, Scotland).

The following POPs were measured: 14 PCB congeners (28, 52, 74, 99, 101, 118, 138, 153, 170, 180, 183, 187, 194, 203), eight OCPs (p,p'-DDE, dichlorodiphenyltrichloroethane [p,p'-DDT], dieldrin, α-endosulfan, heptachlor epoxide, β-hexachlorocyclohexane [β-HCH], γ-HCH, HCB), three PBDE congeners (47, 99, 209), and nine PFAS (perfluorobutyric acid [PFBA], perfluorodecanoic acid [PFDA], perfluorohexanoic acid [PFHxA], perfluorononanoic acid [PFNA], PFOA, perfluoroundecanoic acid [PFUdA], perfluorobutane sulfonate [PFBS], PFHxS and PFOS). However, ten girls of our menstrual characteristics sample had only PFAS measurements due to limited cord blood sample volume. The limit of detection (LOD) ranged from 0.01 (for the majority of these compounds) to 0.3 µg/L (PCB 52). Lipids (TC for total cholesterol, FC for free cholesterol, TG for triglycerides and PL for phospholipids) were measured by an enzymatic method either in plasma samples or serum samples. Total lipids (g/L) were calculated using the following formula: 1.677*(TC-FC) + FC + TG + PL (Akins et al., 1989).

## 2.4. Statistical analyses

POPs with less than 20 % detection rate were not included in the analyses (N = 15 POPs excluded). POPs between 20 and 50 % detection rate were treated dichotomously (detected versus not detected). POPs between 50 and 70 % detection rates were categorised into terciles. POPs with more than 70 % detection rate were categorised into terciles and were also analysed continuously when linearity was confirmed. For these, their concentrations were natural log-transformed and values below the LOD were imputed using maximum likelihood method for left-censored data (five sets) and pooled using Rubin's formula (Rubin, 1996). Our choice of a 70 % detection threshold for imputing via maximum likelihood was informed by Shoari et al. (2015), who showed that this method remains a reliable estimator in these conditions in a dataset with at least 50 observations.

For menstruation characteristics, we performed analyses on the following outcome variables: menarche status for all girls at 12 years old, regularity of menstruations (regularity was self-defined), menstruation duration, severe pain in the lower abdomen or cramps, and severe and prolonged bleeding for girls who had reached menarche.

We studied the associations between each POP and each outcome using a single exposure approach. We used logistic regression models for binary outcomes (menarche status, menstruation regularity, severe pain in the lower abdomen or cramps). Results of logistic regression are presented as odds ratios (OR) and their 95 % confidence interval (95 % CI). We used a Poisson regression model for menstruation duration given its discrete distribution and the absence of over dispersion. Results are presented as percentage changes and their 95 % CI. The main analyses were performed without taking into account lipid concentrations. For severe or prolonged bleedings, regression analyses were not performed due to an insufficient number of events (N = 16). In addition, because cycle length was only collected in girls with regular cycles (N = 64) and due to missing data (N = 15), we did not have a large enough sample size (N = 39) to perform regression analyses on this outcome.

In addition, we performed quantile g-computation to evaluate the mixture effect of the chemicals (of increasing all mixture components by one quartile), by chemical family (OCPs, PCBs, and PFAS) and for all POPs together. Quantile g-computation results are presented as adjusted odds ratio or percentage changes with their 95 % CI.

The selection of potential confounders was determined a priori using a directed acyclic graph (DAG) (Supplementary Fig. 1). We did not adjust on precision variables (predictors of the outcome only) to avoid issues of reduced precision (Schisterman et al., 2009) given our sample size. The following covariates were selected and included in our models: maternal age at delivery (<25, [25–30[, [30–35[, ≥35 years old), pre-pregnancy maternal body mass index (BMI) (<18.5, [18.5–25[, [25–30[, ≥30 kg/m²), prenatal education (lower than baccalaureate, baccalaureate level, higher than baccalaureate), maternal smoking during pregnancy (never or former smoker, smoker in early pregnancy only, smoker at inclusion). Given the small amount of missing data for covariates (<1 %), these were imputed using the mode of each variable.

Sensitivity analyses were performed by additionally adjusting on lipid concentrations when available for all menstruation characteristics. Only girls with total lipids levels ≤ 5 g/L (N = 236) were included because a measurement above this threshold was considered incompatible with cord blood (Spratlen et al., 2020). This analysis was not performed for PFAS due to their lower affinity for lipids (Forsthuber et al., 2020). Total lipids were also natural log-transformed.

All analyses were performed using R (version 4.0.5).

## 3. Results

As described in Table 1, 72.272_2 % of mothers had a university level education. The majority of mothers (58.8 %) were more than 29 years old at inclusion and 60.1 % had at least one previous child. Only three mothers (1.0 %) were considered to have an ethnic background from outside of Europe. Overall, 20.8 % (N = 61) were either smoking at inclusion or had smoked during the first trimester of pregnancy and subsequently quit.

A total of 127 girls (42.9 %) reported the onset of menstruation at 12 years old. Of these, 64 girls (52.0 %) reported regular cycles with a median cycle length of 28 days (inter-quartile range [IQR] 27–30). The median age of menarche among menarcheal girls was 12.1 (IQR 11.0–12.5). The median menstruation duration was 5.0 days (IQR 5.0–6.5). Severe and prolonged bleeding was reported by 12.8 % (N = 16) of girls, while 42.7 % (N = 53) reported experiencing severe lower abdominal pain or cramps. Four girls (1.4 %) were taking oral contraception.

Among the 35 POPs tested, 15 POPs were detected in more than 70 % of samples, with 6 detected in 100 % of samples (Table 2). None of the PBDE congeners were detected in sufficient samples to perform regression analyses.

The majority of POPs were associated (but not statistically significantly) with menarche status at 12 years old (Table 3), especially among PCBs, with an association close to significance for the PCB mixture (OR = 1.29, 95 % CI [0.99; 1.68]). No associations were found between prenatal POPs and regularity of menstruations (Table 4), with no clear pattern regarding the direction of associations. For severe pain in the lower abdomen or cramps, the second tercile of dieldrin exposure was associated with an increased risk (OR = 2.91, 95 % CI [1.13–7.82]), but this association was not present for the third tercile (OR = 0.91, 95 % CI [0.34–2.41]) (Table 5). Similarly, an association was also found between the second tercile of PFNA exposure (OR = 3.13, 95 % CI [1.01–10.43]), but not for the third tercile of exposure (OR = 0.65, 95 % CI [0.18–2.22]). Associations were otherwise inconsistent. POP concentrations generally increased menstrual duration especially for PCBs (Table 6), with the second tercile of HCB exposure being the sole statistically significant association (adjusted percentage change = 25.6 %, 95 % CI [1.2; 56.6]). Associations between the overall mixtures of POPs and showed non-statistically significant longer menstrual duration (1.4 % [−16.1; 22.5]) but a lower risk of severe pain in the lower abdomen or cramps (OR = 0.48, 95 % CI [0.17–1.38]). In sensitivity analyses adjusting on total lipids (for girls with total lipids ≤5 g/L), the results were very similar (Supplementary Tables 1–4).

## 4. Discussion

Using a prospective design, we evaluated the effects of 19 POPs measured in cord blood on menstrual cycle characteristics at 12. Even if we did not observe a statistically significant association between prenatal POP exposure and menarche status at age 12, the majority of POPs, especially PCBs, showed a consistent trend towards an increased
<caption>**Table 1**</caption>

Characteristics of girls who responded to the « Growing and Changing » questionnaire at 12 years old (N = 296).

<table><thead><tr><th>Characteristic</th><th>Categories</th><th>N (%)</th></tr></thead><tbody><tr><td>Maternal age at delivery (years old) (missing data = 0)</td><td>&lt;25<br>[25-29]<br>[30-35]<br>≥35</td><td>11 (3.7)<br>111 (37.5)<br>138 (46.6)<br>36 (12.2)</td></tr><tr><td>Maternal education level at inclusion (missing data = 1)</td><td>Lower than baccalaureate<br>Baccalaureate level<br>Higher than baccalaureate</td><td>28 (9.5)<br>54 (18.3)<br>213 (72.2)</td></tr><tr><td>Maternal ethnicity (missing data = 0)</td><td>At least one parent European<br>Parents not European</td><td>293 (99.0)<br>3 (1.0)</td></tr><tr><td>Parity at inclusion (missing data = 0)</td><td>No previous child<br>At least one child</td><td>118 (39.9)<br>178 (60.1)</td></tr><tr><td>Maternal pre-pregnancy BMI (kg/m²) (missing data = 1)</td><td>&lt;18.5<br>[18.5-25]<br>[25-30]<br>≥30</td><td>19 (6.4)<br>228 (77.3)<br>40 (13.6)<br>8 (2.7)</td></tr><tr><td>Maternal smoking status at inclusion (missing data = 2)</td><td>Never or former smoker<br>Smoker during early pregnancy only<br>Smoker at inclusion</td><td>233 (79.3)<br>37 (12.6)<br>24 (8.2)</td></tr><tr><td>Girl age (median (IQR)) (years) (missing data = 0)</td><td></td><td>12.8 (12.7-12.9)</td></tr><tr><td>Girl BMI (median (IQR)) (kg/m²) (missing data = 28)</td><td></td><td>17.9 (16.6-19.6)</td></tr><tr><td>Girls with menstruations (missing data = 0)</td><td></td><td>127 (42.9)</td></tr><tr><td>Age of menstruations (among girls with menstruations) (median (IQR)) [min-max] (missing data = 8)</td><td></td><td>12.1 (11.0-12.5) [10.0-12.9]</td></tr><tr><td>Regular menstruations (among girls with menstruations) (missing data = 4)</td><td></td><td>64 (52.0)</td></tr><tr><td>Menstrual cycle length (among girls with regular menstruations) (median (IQR)) (days) (missing data = 15)</td><td></td><td>28 (27-30)</td></tr><tr><td>Menstruation duration (among girls with menstruations) (median (IQR)) (days) (missing data = 12)</td><td></td><td>5 (5-6.5)</td></tr><tr><td>Severe and prolonged bleedings (among girls with menstruations) (missing data = 2)</td><td></td><td>16 (12.8)</td></tr><tr><td>Severe pain in the lower abdomen or cramps (among girls with menstruations) (missing data = 3)</td><td></td><td>53 (42.7)</td></tr><tr><td>Girl active smoking (missing data = 5)</td><td></td><td>0 (0.0)</td></tr><tr><td>Girl passive smoking (missing data = 12)</td><td></td><td>93 (32.7)</td></tr><tr><td>Girl contraception (missing data = 17)</td><td></td><td>4 (1.4)</td></tr><tr><td colspan="3">IQR: Inter Quartile Range; BMI: Body Mass Index</td></tr></tbody></table>

probability of menarche at 12 years old and therefore of earlier age at menarche, with a close to significant association for PCBs as a mixture. For severe pain in the lower abdomen or cramps, only the second tercile of dieldrin and of PFNA exposure showed a statistically significant increased risk. For menstruation duration, we found a tendency for POPs to increase menstruation duration, especially for PCBs, and a statistically significant association between the second tercile of HCB and menstruation duration. We did not observe any association for others POPs and other menstrual characteristics. Mixtures of POPs showed non-significant longer menstrual duration, and a non-significant lower risk of severe pain in the lower abdomen or cramps.

Menstrual status and cycle characteristics were in accordance with others population study in girls at the same age (Hoppenbrouwers et al., 2016; Marques et al., 2022; Rigon et al., 2012). We observed a very good concordance for the menarche status between parents and girls (99 %), but only a moderate concordance was observed for regularity, pain and bleedings (between around 64 and 70 %) (data not shown). However, we decided to analyse menstruation characteristics reported by girls and not by parents based on the assumption that girls at 12 years old would be better suited to respond, especially since recall bias regarding menarche is likely very minimal due to the relatively short time between first menstruations and inclusion. Nonetheless, the first menstrual cycles are known to be more variable than at a higher gynaecological age (Harley et al., 2024), which could lead to weaker associations in our study.

The levels of POPs measured in our study were comparable to those reported in the limited number of studies that have measured POPs in cord blood. In the Spanish INMA cohort (1997-98, N = 379 to 660) (Güll-Oumrait et al., 2021), OCP levels were higher, but PFAS concentrations were within a similar range. Conversely, the Belgian EPOPEE cohort (2014-16, N = 58) (Barrea et al., 2024) observed slightly lower POP levels, except for PFOS, which was four times lower than in our study. Additionally, a 2008-09 Chinese mother-child cohort (N = 242) (Wang et al., 2021) reported higher OCP levels but similar PCB concentrations compared to ours. These variations are expected, given the differences in time periods and geographic regions.

To our knowledge, this is the first study investigating the associations between prenatal POPs and menstrual pain or regularity in adolescence. Our results regarding menstruation onset seem to be in line with the available literature studying the effects of prenatal POPs except for PCBs for which we suggest an earlier age at menarche for higher concentration levels. A case-control nested in a British cohort (Avon Longitudinal Study of Parents and Children (ALSPAC)) found no association between prenatal exposure to PCBs, OCPs or PFAS and early menarche status (Christensen et al., 2011; Marks et al., 2021; Namulanda et al., 2016). Kristensen and colleagues found no statistically significant association between prenatal exposure to p,p'-DDE, HCB, a sum of PCBs, and PFAS assessed in maternal serum samples during pregnancy and age at menarche assessed retrospectively in girls at 16 years old or later at 20 years old in a Danish cohort (Kristensen et al., 2013, 2016). Some studies (Harley et al., 2017; Vasiliu et al., 2004) have observed an association between PBDE exposure and a later age at menarche, but we were not able to assess this relation in our study due to low PBDE detection rates. Of note, one study in the United States found that PFNA measured at age 12, and PFHxS measured at age 8 and 12 increased the odds of having reached menarche at 12 years old (Liu et al., 2023) but not when measured in maternal serum during pregnancy despite this measurement being higher on average. None of these studies had evaluated the effects of POPs as a mixture. PFAS in rats exposed at birth have previously been found to suppress kisspeptin fiber intensities, which is crucial in the regulation of puberty (Du et al., 2019), and have been associated with a delayed puberty (Zhang et al., 2021). PFAS have also been shown to affect estrogen and androgen receptors in several animal and in vitro studies (Kjeldsen and Bonefeld-Jørgensen, 2013). PBDEs and PCBs were associated with puberty onset timing (both advance and delay) in female rats exposed during development, possibly through GnRH expression
<table><caption>**Table 2**

Description of prenatal POPs (µg/L) of girls who responded to the « Growing and Changing » questionnaire at 12 years old (N = 296).</caption><thead><tr><th>POPs</th><th>Missing data</th><th>LOD</th><th>N (%) detection rate<sup>a</sup></th><th>min</th><th>25th</th><th>50th</th><th>75th</th><th>max</th></tr></thead><tbody><tr><td colspan="9"><strong>Polychlorinated biphenyls (PCBs)</strong></td></tr><tr><td>PCB 28</td><td>123</td><td>0.05</td><td>0 (0.0)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td></tr><tr><td>PCB 52</td><td>123</td><td>0.30</td><td>0 (0.0)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td></tr><tr><td>PCB 74</td><td>10</td><td>0.02</td><td>45 (15.7)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.088</td></tr><tr><td>PCB 99</td><td>10</td><td>0.02</td><td>47 (16.4)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.062</td></tr><tr><td>PCB 101</td><td>10</td><td>0.01-0.02</td><td>0 (0.0)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td></tr><tr><td>PCB 118</td><td>10</td><td>0.01</td><td>282 (98.6)</td><td>&lt;LOD</td><td>0.020</td><td>0.029</td><td>0.040</td><td>0.140</td></tr><tr><td>PCB 138</td><td>15</td><td>0.01</td><td>281 (100.0)</td><td>0.015</td><td>0.050</td><td>0.068</td><td>0.098</td><td>0.290</td></tr><tr><td>PCB 153</td><td>10</td><td>0.01</td><td>286 (100.0)</td><td>0.028</td><td>0.089</td><td>0.120</td><td>0.170</td><td>0.480</td></tr><tr><td>PCB 170</td><td>10</td><td>0.01</td><td>279 (97.6)</td><td>&lt;LOD</td><td>0.022</td><td>0.029</td><td>0.043</td><td>0.110</td></tr><tr><td>PCB 180</td><td>10</td><td>0.01</td><td>286 (100.0)</td><td>0.014</td><td>0.057</td><td>0.078</td><td>0.110</td><td>0.280</td></tr><tr><td>PCB 183</td><td>10</td><td>0.01</td><td>88 (30.8)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.010</td><td>0.033</td></tr><tr><td>PCB 187</td><td>10</td><td>0.01</td><td>218 (76.2)</td><td>&lt;LOD</td><td>0.010</td><td>0.014</td><td>0.020</td><td>0.080</td></tr><tr><td>PCB 194</td><td>10</td><td>0.01</td><td>117 (40.9)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.013</td><td>0.045</td></tr><tr><td>PCB 203</td><td>10</td><td>0.01</td><td>39 (13.6)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.041</td></tr><tr><td colspan="9"><strong>Organochlorine Pesticides (OCPs)</strong></td></tr><tr><td>p,p'-DDE</td><td>10</td><td>0.02</td><td>269 (94.1)</td><td>&lt;LOD</td><td>0.120</td><td>0.190</td><td>0.310</td><td>3.300</td></tr><tr><td>p,p'-DDT</td><td>10</td><td>0.05</td><td>2 (0.7)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.025</td><td>0.025</td><td>0.066</td></tr><tr><td>Dieldrin</td><td>11</td><td>0.01</td><td>153 (53.7)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.010</td><td>0.020</td><td>0.100</td></tr><tr><td>α-endosulfan</td><td>11</td><td>0.01</td><td>13 (4.6)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.022</td></tr><tr><td>Heptachlor epoxide</td><td>23</td><td>0.02</td><td>79 (28.9)</td><td>&lt;LOD</td><td>0.010</td><td>0.010</td><td>0.010</td><td>0.086</td></tr><tr><td>HCB</td><td>10</td><td>0.02</td><td>260 (90.9)</td><td>&lt;LOD</td><td>0.032</td><td>0.044</td><td>0.057</td><td>0.140</td></tr><tr><td>β-HCH</td><td>10</td><td>0.01</td><td>285 (99.7)</td><td>&lt;LOD</td><td>0.035</td><td>0.051</td><td>0.075</td><td>0.220</td></tr><tr><td>γ-HCH</td><td>10</td><td>0.01</td><td>19 (6.6)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.035</td></tr><tr><td colspan="9"><strong>Polybrominated diphenyl ether (PBDEs)</strong></td></tr><tr><td>PBDE47</td><td>10</td><td>0.01</td><td>53 (18.5)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.190</td></tr><tr><td>PBDE99</td><td>10</td><td>0.01</td><td>32 (11.2)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.210</td></tr><tr><td>PBDE209</td><td>17</td><td>0.02-0.07</td><td>34 (12.2)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.025</td><td>0.860</td></tr><tr><td colspan="9"><strong>Per-polyfluoroalkyl substances (PFAS)</strong></td></tr><tr><td>PFBA</td><td>78</td><td>0.075</td><td>0 (0.0)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td></tr><tr><td>PFBS</td><td>78</td><td>0.066</td><td>5 (2.3)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.251</td></tr><tr><td>PFDA</td><td>99</td><td>0.058</td><td>168 (85.3)</td><td>&lt;LOD</td><td>0.072</td><td>0.099</td><td>0.125</td><td>0.505</td></tr><tr><td>PFHxA</td><td>78</td><td>0.084</td><td>9 (4.1)</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>&lt;LOD</td><td>0.351</td></tr><tr><td>PFHxS</td><td>78</td><td>0.063</td><td>218 (100.0)</td><td>0.064</td><td>0.422</td><td>0.542</td><td>0.686</td><td>1.415</td></tr><tr><td>PFNA</td><td>82</td><td>0.130</td><td>213 (99.5)</td><td>&lt;LOD</td><td>0.248</td><td>0.353</td><td>0.430</td><td>1.541</td></tr><tr><td>PFOA</td><td>78</td><td>0.066</td><td>218 (100.0)</td><td>0.538</td><td>1.657</td><td>2.159</td><td>2.788</td><td>7.362</td></tr><tr><td>PFOS</td><td>78</td><td>0.430</td><td>218 (100.0)</td><td>0.538</td><td>3.430</td><td>4.370</td><td>5.593</td><td>15.556</td></tr><tr><td>PFUdA</td><td>88</td><td>0.048</td><td>157 (75.5)</td><td>&lt;LOD</td><td>0.048</td><td>0.066</td><td>0.088</td><td>0.302</td></tr></tbody></table>

POP: persistent organic pollutant, LOD: Limit Of Detection, p,p'-DDE: Dichlorodiphenyldichloroethylene, p,p'-DDT: Dichlorodiphenyltrichloroethane, HCH: Hexachlorocyclohexane, HCB: Hexachlorobenzene, PFBA: perfluorobutanoic acid, PFBS: Perfluorobutanesulfonic acid, PFDA: perfluorodecanoic acid, PFHxA: perfluorohexanoic acid, PFHxS: Perfluorohexanesulfonic acid, PFNA: perfluoronanoic acid, PFOA: perfluorooctanoic acid, PFOS: Perfluorooctanesulfonic acid, PFUdA: perfluoroundecanoic acid.

disruption (Colciago et al., 2009; Lilienthal et al., 2006).

While we could not assess cycle length due to low sample size, some studies (Kristensen et al., 2013, 2016) have previously reported no statistically significant associations between PFAS or PFOS and cycle length in girls at age 16, and between p,p'-DDE, HCB or a sum of PCBs, and cycle length in women at age 20. Yang et al. (2005) observed shorter menstruation and cycle duration in a subgroup of their population of girls born to mothers exposed to the 1978-79 mass PCBs-PCDFs poisoning from cooking oil in central Taiwan, but generalisation of their results is not possible considering the levels of exposure of the population, the sample size (17 exposed and 16 unexposed) and the age of girls included (between 13 and 19 years old). In our study, we found a non-significant trend between exposure to PCBs, as well as a significant effect of HCB and longer menstruation duration. These results are in agreement with animal studies that have shown longer menstrual duration length for adult monkeys exposed to HCB (Foster et al., 1995), and longer cycle length in rats exposed to Aroclor 1221 (a previously commercialised PCB mixture) in conjunction with a higher expression of Kiss-1 mRNA and AMH mRNA, and lower expression of GnRH mRNA (Hua et al., 2021).

Our study has several limitations. First, due to our longitudinal design (mother-child cohort), our study could be suffering from population selection due to attrition. Sixty percent of the population approached (girls and boys) filled out the questionnaire. The mothers of the girls included in this study were significantly more educated (72.2 %

with more than baccalaureate vs 62.4 %), and smoked less than those of the original cohort (79.3 % not smoker or former smoker vs 71.9 %) (data not shown). However, we have no data indicating this selection could bias the relationship between POPs and menstrual characteristics. Second, lipophilic POPs were measured in three different batches. However, we believe this difference has little influence on the determination of POPs since the batches were measured in the same laboratory with very good internal reference. Moreover, we decided not to adjust on batch since a different in batches could not be considered as confounder as POP measurements were determined independently of menstrual characteristics. Third, menstrual cycle characteristics may be subject to recall bias (cycle length and menstruation duration) or influenced by subjective reporting (regularity, severe pain and cramps); however, as these potential inaccuracies are unlikely to differ by levels of POP exposure, any resulting misclassification is expected to be non-differential, which would likely bias observed associations toward the null. Fourth, although consistent with similar mother-child cohorts, our sample size was limited by the number of available POP measurements and could be affected by a lack of statistical power.

Our study also has several strengths, including its prospective design and the measurement of a wide range of POPs at birth, which likely serves as a reliable proxy for prenatal exposure due to the long half-life of these chemicals. Additionally, POP measurements were conducted using cord blood, which is regarded as a more accurate indicator of prenatal exposure than maternal serum. Finally, we employed both

<footnote>a POPs with detection frequencies <20 % were excluded from the regression analysis due to low detection.</footnote>
<caption>**Table 3**</caption>

Associations between prenatal exposure to POPs and menarche status at the 12 years old follow-up (N = 296).

<table><caption>**Table 3 (continued)**</caption><thead><tr><th>POPs</th><th>Concentration levels (µg/L)</th><th>No menarche (N = 169) N (%)</th><th>Menarche (N = 127) N (%)</th><th>Adjusted OR (95% CI)<sup>a</sup></th></tr></thead><tbody><tr><td colspan="5"><strong>Polychlorinated biphenyls (PCBs)</strong></td></tr><tr><td>PCB 118</td><td>Tercile 1 (&lt;0.023)<br/>Tercile 2 ([0.023-0.036])<br/>Tercile 3 (&gt;0.036)<br/>Continuous<sup>b</sup></td><td>60 (36.4)<br/>58 (35.2)<br/>47 (28.5)</td><td>39 (32.2)<br/>39 (32.2)<br/>43 (35.5)</td><td>ref.<br/>1.01 (0.56; 1.84)<br/>1.35 (0.73; 2.52)<br/>1.17 (0.72; 1.91)</td></tr><tr><td>PCB 138</td><td>Tercile 1 (&lt;0.055)<br/>Tercile 2 ([0.055-0.083])<br/>Tercile 3 (&gt;0.083)<br/>Continuous<sup>b</sup></td><td>57 (35.4)<br/>57 (35.4)<br/>47 (29.2)</td><td>38 (31.7)<br/>37 (30.8)<br/>45 (37.5)</td><td>ref.<br/>0.95 (0.51; 1.73)<br/>1.43 (0.77; 2.68)<br/>1.55 (0.94; 2.59)</td></tr><tr><td>PCB 153</td><td>Tercile 1 (&lt;0.100)<br/>Tercile 2 ([0.100-0.150])<br/>Tercile 3 (&ge;0.150)<br/>Continuous<sup>b</sup></td><td>54 (32.7)<br/>58 (35.2)<br/>53 (32.1)</td><td>34 (28.1)<br/>38 (31.4)<br/>49 (40.5)</td><td>ref.<br/>1.01 (0.54; 1.89)<br/>1.55 (0.81; 2.97)<br/>1.46 (0.87; 2.49)</td></tr><tr><td>PCB 170</td><td>Tercile 1 (&lt;0.024)<br/>Tercile 2 ([0.024-0.037])<br/>Tercile 3 (&ge;0.037)<br/>Continuous<sup>b</sup></td><td>58 (35.2)<br/>51 (30.9)<br/>56 (33.9)</td><td>34 (28.1)<br/>43 (35.5)<br/>44 (36.4)</td><td>ref.<br/>1.46 (0.79; 2.71)<br/>1.55 (0.80; 3.02)<br/>1.41 (0.85; 2.35)</td></tr><tr><td>PCB 180</td><td>Tercile 1 (&lt;0.064)<br/>Tercile 2 ([0.064-0.095])<br/>Tercile 3 (&ge;0.095)<br/>Continuous<sup>b</sup></td><td>58 (35.2)<br/>51 (30.9)<br/>56 (33.9)</td><td>37 (30.6)<br/>43 (35.5)<br/>41 (33.9)</td><td>ref.<br/>1.31 (0.71; 2.43)<br/>1.30 (0.67; 2.53)<br/>1.42 (0.85; 2.39)</td></tr><tr><td>PCB 183</td><td>&lt;LOD<br/>&ge;LOD</td><td>120 (72.7)<br/>45 (27.3)</td><td>78 (64.5)<br/>43 (35.5)</td><td>ref.<br/>1.56 (0.91; 2.67)</td></tr><tr><td>PCB 187</td><td>Tercile 1 (&lt;0.010)<br/>Tercile 2 ([0.010-0.017])<br/>Tercile 3 (&gt;0.017)<br/>Continuous<sup>b</sup></td><td>60 (36.4)<br/>55 (33.3)<br/>50 (30.3)</td><td>38 (31.4)<br/>42 (34.7)<br/>41 (33.9)</td><td>ref.<br/>1.34 (0.73; 2.48)<br/>1.46 (0.77; 2.81)<br/>1.21 (0.72; 2.05)</td></tr><tr><td>PCB 194</td><td>&lt;LOD<br/>&ge;LOD</td><td>99 (60.0)<br/>66 (40.0)</td><td>70 (57.9)<br/>51 (42.1)</td><td>ref.<br/>1.30 (0.76; 2.23)<br/>1.29 (0.99; 1.68)</td></tr><tr><td colspan="5"><strong>Organochlorine Pesticides (OCPs)</strong></td></tr><tr><td>p,p'-DDE</td><td>Tercile 1 (&lt;0.15)<br/>Tercile 2 ([0.15-0.25])<br/>Tercile 3 (&ge;0.25)<br/>Continuous<sup>b</sup></td><td>55 (33.3)<br/>55 (33.3)<br/>55 (33.3)</td><td>37 (30.6)<br/>39 (32.2)<br/>45 (37.2)</td><td>ref.<br/>0.95 (0.52; 1.75)<br/>1.10 (0.60; 2.01)<br/>1.18 (0.89; 1.57)</td></tr><tr><td>Dieldrin</td><td>Tercile 1 (&lt;0.01)<br/>Tercile 2 ([0.01-0.020])<br/>Tercile 3 (&ge;0.020)</td><td>72 (43.6)<br/>44 (26.7)<br/>49 (29.7)</td><td>60 (50.0)<br/>30 (25.0)<br/>30 (25.0)</td><td>ref.<br/>0.81 (0.44; 1.45)<br/>0.69 (0.38; 1.24)</td></tr></tbody></table>

<table><thead><tr><th>POPs</th><th>Concentration levels (µg/L)</th><th>No menarche (N = 169) N (%)</th><th>Menarche (N = 127) N (%)</th><th>Adjusted OR (95% CI)<sup>a</sup></th></tr></thead><tbody><tr><td>Heptachlor epoxide</td><td>&lt;LOD<br/>&ge;LOD</td><td>120 (75.0)<br/>40 (25.0)</td><td>74 (65.5)<br/>39 (34.5)</td><td>ref.<br/>1.48 (0.86; 2.55)</td></tr><tr><td rowspan="4">β-HCH</td><td>Tercile 1 (&lt;0.039)</td><td>60 (36.4)</td><td>36 (29.8)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.039-0.064])</td><td>55 (33.3)</td><td>39 (32.2)</td><td>1.18 (0.65; 2.16)</td></tr><tr><td>Tercile 3 (&ge;0.064)</td><td>50 (30.3)</td><td>46 (38.0)</td><td>1.50 (0.82; 2.78)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.29 (0.84; 1.97)</td></tr><tr><td rowspan="4">HCB</td><td>Tercile 1 (&lt;0.036)</td><td>61 (37.0)</td><td>35 (28.9)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.036-0.053])</td><td>49 (29.7)</td><td>45 (37.2)</td><td>1.63 (0.90; 2.97)</td></tr><tr><td>Tercile 3 (&ge;0.053)</td><td>55 (33.3)</td><td>41 (33.9)</td><td>1.26 (0.69; 2.32)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.33 (0.77; 2.30)</td></tr><tr><td>OCP mixture<sup>c</sup></td><td></td><td></td><td></td><td>1.15 (0.88; 1.51)</td></tr><tr><td colspan="5"><strong>Per-polyfluoroalkyl substances (PFAS)</strong></td></tr><tr><td rowspan="4">PFDA</td><td>Tercile 1 (&lt;0.082)</td><td>38 (33.0)</td><td>28 (34.1)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.082-0.114])</td><td>39 (33.9)</td><td>28 (34.1)</td><td>0.99 (0.47; 2.06)</td></tr><tr><td>Tercile 3 (&gt;0.114)</td><td>38 (33.0)</td><td>26 (31.7)</td><td>1.04 (0.49; 2.20)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.32 (0.62; 2.83)</td></tr><tr><td rowspan="4">PFNA</td><td>Tercile 1 (&lt;0.294)</td><td>41 (32.5)</td><td>30 (34.1)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.294-0.409])</td><td>39 (31.0)</td><td>32 (36.4)</td><td>1.19 (0.58; 2.42)</td></tr><tr><td>Tercile 3 (&ge;0.409)</td><td>46 (36.5)</td><td>26 (29.5)</td><td>0.82 (0.40; 1.67)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.30 (0.63; 2.69)</td></tr><tr><td rowspan="4">PFOA</td><td>Tercile 1 (&lt;1.080)</td><td>43 (33.6)</td><td>30 (33.3)</td><td>ref.</td></tr><tr><td>Tercile 2 ([1.080-2.557])</td><td>42 (32.8)</td><td>30 (33.3)</td><td>1.03 (0.51; 2.09)</td></tr><tr><td>Tercile 3 (&ge;2.557)</td><td>43 (33.6)</td><td>30 (33.3)</td><td>1.04 (0.51; 2.13)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.30 (0.63; 2.69)</td></tr><tr><td rowspan="4">PFUdA</td><td>Tercile 1 (&lt;0.051)</td><td>41 (33.9)</td><td>29 (33.3)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.051-0.080])</td><td>36 (29.8)</td><td>33 (37.9)</td><td>1.68 (0.80; 3.59)</td></tr><tr><td>Tercile 3 (&gt;0.080)</td><td>44 (36.4)</td><td>25 (28.7)</td><td>1.08 (0.51; 2.33)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.52 (0.74; 3.10)</td></tr><tr><td rowspan="4">PFHxS</td><td>Tercile 1 (&lt;0.456)</td><td>42 (32.8)</td><td>29 (32.2)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.456-0.647])</td><td>44 (34.4)</td><td>30 (33.3)</td><td>0.78 (0.38; 1.58)</td></tr><tr><td>Tercile 3 (&ge;0.647)</td><td>42 (32.8)</td><td>31 (34.4)</td><td>1.02 (0.50; 2.08)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.40 (0.72; 2.77)</td></tr><tr><td rowspan="4">PFOS</td><td>Tercile 1 (&lt;3.736)</td><td>47 (36.7)</td><td>26 (28.9)</td><td>ref.</td></tr><tr><td>Tercile 2 ([3.736-5.074])</td><td>44 (34.4)</td><td>28 (31.1)</td><td>0.88 (0.42; 1.82)</td></tr><tr><td>Tercile 3 (&ge;5.074)</td><td>37 (28.9)</td><td>36 (40.0)</td><td>1.47 (0.73; 2.99)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.66 (0.84; 3.41)</td></tr><tr><td>PFAS mixture</td><td></td><td></td><td></td><td>1.15 (0.78; 1.70)</td></tr><tr><td>Σ POP mixture</td><td></td><td></td><td></td><td>1.57 (0.88; 2.79)</td></tr></tbody></table>
POP: persistent organic pollutant; Ref: reference; LOD: Limit Of Detection, p,p'-DDE: Dichlorodiphenyldichloroethylene, p,p'-DDT: Dichlorodiphenyltrichloroethane, HCH: Hexachlorocyclohexane, HCB: Hexachlorobenzene, PFBA: perfluorobutanoic acid, PFBS: Perfluorobutanesulfonic acid, PFDA: perfluorodecanoic acid, PFHxA: perfluorohexanoic acid, PFHxS: Perfluorohexanesulfonic acid, PFNA: perfluorononanoic acid, PFOA: perfluorooctanoic acid, PFOS: Perfluorooctanesulfonic acid, PFUdA: perfluoroundecanoic acid.

a Adjusted on maternal age at delivery, maternal pre-pregnancy body mass index, prenatal smoking status, prenatal maternal education level.

b OR are given for natural log-transformed POP analysed continuously when linearity was accepted.

c Effect for one quartile increase of all included chemicals using quantile g-computation.

single-exposure and complementary mixture analysis methods to assess the impact of pollutants on menstrual characteristics.

## 5. Conclusion

Our results suggested a possible impact of prenatal exposure to POPs on some menstruation characteristics of girls at age 12, including a lower age at menarche, but these need to be further confirmed at a later age and in prospective studies with a larger sample size, possibly through the combined analysis of several mother-child cohorts. This topic is of particular interest in a context where several studies have observed secular trends toward earlier puberty in girls, though the role of chemicals such as POPs in this decrease remains unknown. In addition, although POP levels appear to be decreasing in parts of Europe, this trend is not consistent across all populations. PFAS, in particular, remain highly persistent in the environment, and newer short-chain compounds are continuing to emerge. For these reasons, the health effects of this class of chemicals warrant close and ongoing monitoring. Finally, analyses examining the potential mediating role of steroid hormones in the relationship between POP exposure and puberty-related outcomes would be highly relevant.

## CRediT authorship contribution statement

**Maximilien Génard-Walton:** Writing – review & editing, Writing – original draft, Methodology, Formal analysis. **Elisabeth Angot:** Writing – review & editing, Methodology, Formal analysis. **Charline Warembourg:** Writing – review & editing, Methodology. **Christine Monfort:** Writing – review & editing, Investigation, Data curation. **Florence Rouget:** Writing – review & editing, Supervision, Methodology, Conceptualization. **Nathalie Costet:** Writing – review & editing, Methodology, Conceptualization. **Hélene Tillault:** Writing – review & editing, Methodology, Data curation. **Fabrice Lainé:** Writing – review & editing, Data curation. **Éric Gaudreau:** Writing – review & editing, Data curation. **Sylvaine Cordier:** Writing – review & editing, Methodology, Conceptualization. **Marina Kvaskoff:** Writing – review & editing, Methodology. **Cécile Chevrier:** Writing – review & editing, Supervision, Methodology, Investigation, Funding acquisition, Conceptualization. **Ronan Garlantézec:** Writing – review & editing, Supervision, Methodology, Funding acquisition, Conceptualization.

## Ethics approval

The study was approved by the relevant ethical committee for studies involving human subjects (Comité de Protection des Personnes), and the French national security agency for medicine and health products (ANSM). All participants provided written informed consent for their participation in the study.

## Funding sources

The PELAGIE cohort has been funded since its inception by the

<caption>**Table 4**</caption>

Associations between prenatal exposure to POPs and menstrual regularity at the 12 years old follow-up (N = 296).

<table><thead><tr><th></th><th>Concentration levels (µg/L)</th><th>Unregular menstruations (N = 59) N (%)</th><th>Regular menstruations (N = 64) N (%)</th><th>Adjusted OR<sup>a</sup> (CI95 %)</th></tr></thead><tbody><tr><td rowspan="4">PCB 118</td><td>Tercile 1 (&lt;0.023)</td><td>18 (32.1)</td><td>20 (32.8)</td><td>ref.</td></tr><tr><td>Tercile 2 (&gt;0.023–0.036)</td><td>21 (37.5)</td><td>17 (27.9)</td><td>1.37 (0.52; 3.66)</td></tr><tr><td>Tercile 3 (&gt;0.036)</td><td>17 (30.4)</td><td>24 (39.3)</td><td>0.82 (0.31; 2.18)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>0.98 (0.45; 2.15)</td></tr><tr><td rowspan="4">PCB 138</td><td>Tercile 1 (&lt;0.055)</td><td>18 (32.1)</td><td>19 (31.7)</td><td>ref.</td></tr><tr><td>Tercile 2 (&gt;0.055–0.083)</td><td>17 (30.4)</td><td>19 (31.7)</td><td>0.78 (0.28; 2.14)</td></tr><tr><td>Tercile 3 (&gt;0.083)</td><td>21 (37.5)</td><td>22 (36.7)</td><td>0.95 (0.35; 2.55)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.07 (0.47; 2.44)</td></tr><tr><td rowspan="4">PCB 153</td><td>Tercile 1 (&lt;0.100)</td><td>16 (28.6)</td><td>17 (27.9)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.100–0.150])</td><td>16 (28.6)</td><td>21 (34.4)</td><td>0.75 (0.27; 2.07)</td></tr><tr><td>Tercile 3 (&gt;0.150)</td><td>24 (42.9)</td><td>23 (37.7)</td><td>1.1 (0.40; 3.04)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.03 (0.44; 2.41)</td></tr><tr><td rowspan="4">PCB 170</td><td>Tercile 1 (&lt;0.024)</td><td>14 (25.0)</td><td>19 (31.1)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.024–0.037])</td><td>21 (37.5)</td><td>21 (34.4)</td><td>1.18 (0.44; 3.16)</td></tr><tr><td>Tercile 3 (&gt;0.037)</td><td>21 (37.5)</td><td>21 (34.4)</td><td>1.35 (0.48; 3.83)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.17 (0.51; 2.70)</td></tr><tr><td rowspan="4">PCB 180</td><td>Tercile 1 (&lt;0.064)</td><td>15 (26.8)</td><td>21 (34.4)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.064–0.095])</td><td>21 (37.5)</td><td>21 (34.4)</td><td>1.43 (0.55; 3.77)</td></tr><tr><td>Tercile 3 (&gt;0.095)</td><td>20 (35.7)</td><td>19 (31.1)</td><td>1.61 (0.57; 4.65)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.20 (0.50; 2.87)</td></tr><tr><td rowspan="2">PCB 183</td><td>&lt;LOD</td><td>35 (62.5)</td><td>41 (67.2)</td><td>ref.</td></tr><tr><td>&ge;LOD</td><td>21 (37.5)</td><td>20 (32.8)</td><td>1.22 (0.54; 2.77)</td></tr><tr><td rowspan="4">PCB 187</td><td>Tercile 1 (&lt;0.010)</td><td>16 (28.6)</td><td>21 (34.4)</td><td>ref.</td></tr><tr><td>Tercile 2 (&gt;0.010–0.017)</td><td>21 (37.5)</td><td>20 (32.8)</td><td>1.36 (0.52; 3.61)</td></tr><tr><td>Tercile 3 (&gt;0.017)</td><td>19 (33.9)</td><td>20 (32.8)</td><td>1.17 (0.43; 3.22)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>0.95 (0.39; 2.35)</td></tr><tr><td>PCB 194</td><td>&lt;LOD</td><td>33 (58.9)</td><td>36 (59.0)</td><td>ref.</td></tr></tbody></table>

(continued on next page)
<table><caption>Table 4 (continued)</caption><thead><tr><th></th><th>Concentration levels (µg/L)</th><th>Unregular menstruations (N = 59) N (%)</th><th>Regular menstruations (N = 64) N (%)</th><th>Adjusted OR<sup>a</sup> (CI95 %)</th></tr></thead><tbody><tr><td></td><td>≥LOD</td><td>23 (41.1)</td><td>25 (41.0)</td><td>0.95 (0.41; 2.20)</td></tr><tr><td>PCB mixture<sup>c</sup></td><td></td><td></td><td></td><td>1.06 (0.70; 1.62)</td></tr><tr><td>p,p'-DDE</td><td>Tercile 1 (&lt;0.15)</td><td>19 (33.9)</td><td>18 (29.5)</td><td>ref.</td></tr><tr><td></td><td>Tercile 2 ([0.15-0.25])</td><td>16 (28.6)</td><td>21 (34.4)</td><td>0.75 (0.28; 1.98)</td></tr><tr><td></td><td>Tercile 3 (≥0.25)</td><td>21 (37.5)</td><td>22 (36.1)</td><td>0.96 (0.37; 2.47)</td></tr><tr><td></td><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.03 (0.64; 1.66)</td></tr><tr><td>Dieldrin</td><td>Tercile 1 (&lt;0.01)</td><td>28 (50.9)</td><td>31 (50.8)</td><td>ref.</td></tr><tr><td></td><td>Tercile 2 ([0.01-0.020])</td><td>11 (20.0)</td><td>16 (26.2)</td><td>0.88 (0.33; 2.34)</td></tr><tr><td></td><td>Tercile 3 (≥0.020)</td><td>16 (29.1)</td><td>14 (23.0)</td><td>1.40 (0.54; 3.67)</td></tr><tr><td>Heptachlor epoxide</td><td>&lt;LOD</td><td>31 (60.8)</td><td>41 (69.5)</td><td>ref.</td></tr><tr><td></td><td>≥LOD</td><td>20 (39.2)</td><td>18 (30.5)</td><td>1.49 (0.65; 3.45)</td></tr><tr><td>B-HCH</td><td>Tercile 1 (≤0.039)</td><td>16 (28.6)</td><td>19 (31.1)</td><td>ref.</td></tr><tr><td></td><td>Tercile 2 [] (0.039-0.064[])</td><td>17 (30.4)</td><td>21 (34.4)</td><td>1.00 (0.38; 2.66)</td></tr><tr><td></td><td>Tercile 3 (≥0.064)</td><td>23 (41.1)</td><td>21 (34.4)</td><td>1.27 (0.47; 3.44)</td></tr><tr><td>HCB</td><td>Tercile 1 (≤0.036)</td><td>16 (28.6)</td><td>18 (29.5)</td><td>ref.</td></tr><tr><td></td><td>Tercile 2 [] (0.036-0.053[])</td><td>22 (39.3)</td><td>21 (34.4)</td><td>0.96 (0.37; 2.48)</td></tr><tr><td></td><td>Tercile 3 (≥0.053)</td><td>18 (32.1)</td><td>22 (36.1)</td><td>0.83 (0.30; 2.30)</td></tr><tr><td></td><td>Continuous<sup>b</sup></td><td></td><td></td><td>0.80 (0.31; 2.07)</td></tr><tr><td>OCP mixture</td><td></td><td></td><td></td><td>1.03 (0.67; 1.58)</td></tr><tr><td>PFDA</td><td>Tercile 1 (&lt;0.082)</td><td>14 (35.0)</td><td>13 (33.3)</td><td>ref.</td></tr><tr><td></td><td>Tercile 2 ([0.082-0.114])</td><td>16 (40.0)</td><td>12 (30.8)</td><td>1.39 (0.44; 4.58)</td></tr><tr><td></td><td>Tercile 3 (&gt;0.114)</td><td>10 (25.0)</td><td>14 (35.9)</td><td>0.76 (0.24; 2.44)</td></tr><tr><td></td><td>Continuous<sup>b</sup></td><td></td><td></td><td>0.53 (0.16; 1.79)</td></tr><tr><td>PFNA</td><td>Tercile 1 (≤0.294)</td><td>14 (34.1)</td><td>15 (34.1)</td><td>ref.</td></tr><tr><td></td><td>Tercile 2 ([0.294-0.409[])</td><td>13 (31.7)</td><td>18 (40.9)</td><td>0.88 (0.30; 2.60)</td></tr><tr><td></td><td>Tercile 3 (≥0.409)</td><td>14 (34.1)</td><td>11 (25.0)</td><td>1.72 (0.55; 5.59)</td></tr><tr><td></td><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.01 (0.29; 3.53)</td></tr></tbody></table>

<table><caption>Table 4 (continued)</caption><thead><tr><th></th><th>Concentration levels (µg/L)</th><th>Unregular menstruations (N = 59) N (%)</th><th>Regular menstruations (N = 64) N (%)</th><th>Adjusted OR<sup>a</sup> (CI95 %)</th></tr></thead><tbody><tr><td>PFOA</td><td>Tercile 1 (&lt;1.808)</td><td>13 (31.0)</td><td>17 (37.8)</td><td>ref.</td></tr><tr><td></td><td>Tercile 2 ([1.808-2.557[])</td><td>13 (31.0)</td><td>14 (31.1)</td><td>1.42 (0.48; 4.35)</td></tr><tr><td></td><td>Tercile 3 (≥2.557)</td><td>16 (38.1)</td><td>14 (31.1)</td><td>1.90 (0.64; 5.79)</td></tr><tr><td>PFUdA</td><td>Tercile 1 (≤0.051)</td><td>16 (39.0)</td><td>12 (27.9)</td><td>ref.</td></tr><tr><td></td><td>Tercile 2 [] (0.051-0.080[])</td><td>16 (39.0)</td><td>17 (39.5)</td><td>0.83 (0.28; 2.51)</td></tr><tr><td></td><td>Tercile 3 (&gt;0.080)</td><td>9 (22.0)</td><td>14 (32.6)</td><td>0.50 (0.15; 1.58)</td></tr><tr><td></td><td>Continuous<sup>b</sup></td><td></td><td></td><td>0.44 (0.14; 1.36)</td></tr><tr><td>PFHxS</td><td>Tercile 1 (&lt;0.456)</td><td>13 (31.0)</td><td>15 (33.3)</td><td>ref.</td></tr><tr><td></td><td>Tercile 2 ([0.456-0.647[])</td><td>12 (28.6)</td><td>16 (35.6)</td><td>1.06 (0.34; 3.37)</td></tr><tr><td></td><td>Tercile 3 (≥0.647)</td><td>17 (40.5)</td><td>14 (31.1)</td><td>1.68 (0.58; 5.05)</td></tr><tr><td></td><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.62 (0.55; 4.97)</td></tr><tr><td>PFOS</td><td>Tercile 1 (&lt;3.736)</td><td>11 (26.2)</td><td>14 (31.1)</td><td>ref.</td></tr><tr><td></td><td>Tercile 2 ([3.736-5.074[])</td><td>15 (35.7)</td><td>12 (26.7)</td><td>1.72 (0.53; 5.74)</td></tr><tr><td></td><td>Tercile 3 (≥5.074)</td><td>16 (38.1)</td><td>19 (42.2)</td><td>1.16 (0.39; 3.52)</td></tr><tr><td>PFAS mixture</td><td></td><td></td><td></td><td>1.14 (0.62; 2.09)</td></tr><tr><td>ΣPOP mixture</td><td></td><td></td><td></td><td>1.14 (0.22; 2.40)</td></tr></tbody></table>

POP: persistent organic pollutant; Ref: reference; LOD: Limit Of Detection, p,p'-DDE: Dichlorodiphenyldichloroethylene, p,p'-DDT: Dichlorodiphenyltrichloroethane, HCH: Hexachlorocyclohexane, HCB: Hexachlorobenzene, PFBA: perfluorobutanoic acid, PFBS: Perfluorobutanesulfonic acid, PFDA: perfluorodecanoic acid, PFHxA: perfluorohexanoic acid, PFHxS: Perfluorohexanesulfonic acid, PFNA: perfluorononanoic acid, PFOA: perfluorooctanoic acid, PFOS: Perfluorooctanesulfonic acid, PFUdA: perfluoroundecanoic acid.

a Adjusted on maternal age at delivery, maternal pre-pregnancy body mass index, prenatal smoking status, prenatal maternal education level.

b OR are given for natural log-transformed POP analysed continuously when linearity was accepted.

c Effect for one quartile increase of all included chemicals using quantile g-computation.

Institut national de la santé et de la recherche médicale (Inserm), the French Ministries of Health (2003–2004), Labour (2002–2003), and Research (ATC 2003–2004), the French National Institute for Public Health Surveillance (InVS, 2002–2006), the National Agency for Research (ANR, 2005–2008, 2010–2012, 2015–2019), the French Agency for Environmental Health Safety (Afsset/ANSES, 2007–2009, 2009–2012, 2019–2023), the French Agency for Drug Safety (2013–2017), the Fondation de France (2014–2017, 2015–2018, 2017–2020, 2019–2021, 2021–2024), the French Ministry of Ecology (PNRPE 2014–2016), the Research Institute of Public Health (IResP)
<caption>**Table 5**</caption>

Associations between prenatal exposure to POPs and severe pain in the lower abdomen or cramps at the 12 years old follow-up (N = 124).

<table><caption>**Table 5 (continued)**</caption><thead><tr><th>POPs</th><th>Concentration levels (µg/L)</th><th>No pain (N = 71) N (%)</th><th>Pain (N = 53) N (%)</th><th>Adjusted OR <sup>a</sup> (95 % CI)</th></tr></thead><tbody><tr><td colspan="5"><strong>Polychlorinated biphenyls (PCBs)</strong></td></tr><tr><td rowspan="4">PCB 118</td><td>Tercile 1 (&le;0.023)</td><td>21 (31.3)</td><td>16 (31.4)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.023-0.036])</td><td>23 (34.3)</td><td>16 (31.4)</td><td>1.03 (0.39; 2.74)</td></tr><tr><td>Tercile 3 (&gt;0.036)</td><td>23 (34.3)</td><td>19 (37.3)</td><td>1.34 (0.52; 3.55)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>0.84 (0.38; 1.82)</td></tr><tr><td rowspan="4">PCB 138</td><td>Tercile 1 (&le;0.055)</td><td>20 (30.3)</td><td>17 (33.3)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.055-0.083])</td><td>19 (28.8)</td><td>18 (35.3)</td><td>1.33 (0.50; 3.58)</td></tr><tr><td>Tercile 3 (&gt;0.083)</td><td>27 (40.9)</td><td>16 (31.4)</td><td>0.83 (0.31; 2.20)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>0.86 (0.39; 1.89)</td></tr><tr><td rowspan="4">PCB 153</td><td>Tercile 1 (&lt;0.100)</td><td>17 (25.4)</td><td>17 (33.3)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.100-0.150])</td><td>21 (31.3)</td><td>16 (31.4)</td><td>0.85 (0.32; 2.26)</td></tr><tr><td>Tercile 3 (&ge;0.150)</td><td>29 (43.3)</td><td>18 (35.3)</td><td>0.72 (0.27; 1.91)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>0.77 (0.33; 1.75)</td></tr><tr><td rowspan="4">PCB 170</td><td>Tercile 1 (&lt;0.024)</td><td>17 (25.4)</td><td>17 (33.3)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.024-0.037])</td><td>24 (35.8)</td><td>18 (35.3)</td><td>0.82 (0.32; 2.13)</td></tr><tr><td>Tercile 3 (&ge;0.037)</td><td>26 (38.8)</td><td>16 (31.4)</td><td>0.69 (0.25; 1.88)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>0.75 (0.33; 1.69)</td></tr><tr><td rowspan="4">PCB 180</td><td>Tercile 1 (&lt;0.064)</td><td>18 (26.9)</td><td>18 (35.3)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.064-0.095])</td><td>27 (40.3)</td><td>16 (31.4)</td><td>0.66 (0.26; 1.68)</td></tr><tr><td>Tercile 3 (&ge;0.095)</td><td>22 (32.8)</td><td>17 (33.3)</td><td>0.92 (0.33; 2.53)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>0.76 (0.32; 1.77)</td></tr><tr><td rowspan="4">PCB 183</td><td>&lt;LOD</td><td>41 (61.2)</td><td>36 (70.6)</td><td>ref.</td></tr><tr><td>&ge;LOD</td><td>26 (38.8)</td><td>15 (29.4)</td><td>0.71 (0.31; 1.57)</td></tr><tr><td rowspan="4">PCB 187</td><td>Tercile 1 (&le;0.010)</td><td>22 (32.8)</td><td>15 (29.4)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.010-0.017])</td><td>22 (32.8)</td><td>20 (39.2)</td><td>1.53 (0.59; 4.06)</td></tr><tr><td>Tercile 3 (&gt;0.017)</td><td>23 (34.3)</td><td>16 (31.4)</td><td>1.23 (0.46; 3.38)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.00 (0.44; 2.27)</td></tr><tr><td rowspan="2">PCB 194</td><td>&lt;LOD</td><td>39 (58.2)</td><td>30 (58.8)</td><td>ref.</td></tr><tr><td>&ge;LOD</td><td>28 (41.8)</td><td>21 (41.2)</td><td>1.09 (0.47; 2.50)</td></tr><tr><td colspan="4">PCB mixture<sup>c</sup></td><td>0.93 (0.62; 1.38)</td></tr><tr><td colspan="5"><strong>Organochlorine Pesticides (OCPs)</strong></td></tr><tr><td rowspan="4">p,p'-DDE</td><td>Tercile 1 (&lt;0.15)</td><td>21 (31.3)</td><td>15 (29.4)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.15-0.25])</td><td>22 (32.8)</td><td>16 (31.4)</td><td>1.12 (0.43; 2.95)</td></tr><tr><td>Tercile 3 (&ge;0.25)</td><td>24 (35.8)</td><td>20 (39.2)</td><td>1.39 (0.55; 3.62)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>1.20 (0.75; 1.93)</td></tr><tr><td rowspan="4">Dieldrin</td><td>Tercile 1 (&lt;0.01)</td><td>36 (53.7)</td><td>22 (44.0)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.01-0.020])</td><td>11 (16.4)</td><td>18 (36.0)</td><td>2.91 (1.13; 7.82)</td></tr><tr><td>Tercile 3 (&ge;0.020)</td><td>20 (29.9)</td><td>10 (20.0)</td><td>0.91 (0.34; 2.41)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td></td></tr><tr><td rowspan="2">Heptachlor epoxide</td><td>&lt;LOD</td><td>39 (63.9)</td><td>33 (67.3)</td><td>ref.</td></tr><tr><td>&ge;LOD</td><td>22 (36.1)</td><td>16 (32.7)</td><td>0.86 (0.37; 1.93)</td></tr><tr><td rowspan="2">β-HCH</td><td>Tercile 1 (&le;0.039)</td><td>19 (28.4)</td><td>17 (33.3)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.039-0.064])</td><td>17 (25.4)</td><td>20 (39.2)</td><td>1.42 (0.55; 3.70)</td></tr></tbody></table>

<table><thead><tr><th>POPs</th><th>Concentration levels (µg/L)</th><th>No pain (N = 71) N (%)</th><th>Pain (N = 53) N (%)</th><th>Adjusted OR <sup>a</sup> (95 % CI)</th></tr></thead><tbody><tr><td rowspan="4">HCB</td><td>Tercile 3 (&ge;0.064)</td><td>31 (46.3)</td><td>14 (27.5)</td><td>0.60 (0.22; 1.59)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>0.71 (0.37; 1.37)</td></tr><tr><td>Tercile 1 (&le;0.036)</td><td>18 (26.9)</td><td>16 (31.4)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.036-0.053])</td><td>24 (35.8)</td><td>19 (37.3)</td><td>0.98 (0.38; 2.50)</td></tr><tr><td rowspan="4">OCP mixture</td><td>Tercile 3 (&ge;0.053)</td><td>25 (37.3)</td><td>16 (31.4)</td><td>0.91 (0.34; 2.46)</td></tr><tr><td>Continuous<sup>b</sup></td><td></td><td></td><td>0.88 (0.35; 2.22)</td></tr><tr><td></td><td></td><td></td><td>0.91 (0.59; 1.38)</td></tr><tr><td>Per-polyfluoroalkyl substances (PFAS)</td><td></td><td></td><td></td><td></td></tr><tr><td rowspan="3">PFDA</td><td>Tercile 1 (&lt;0.082)</td><td>16 (34.8)</td><td>10 (29.4)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.082-0.114])</td><td>15 (32.6)</td><td>13 (38.2)</td><td>0.93 (0.27; 3.16)</td></tr><tr><td>Tercile 3 (&gt;0.114)</td><td>15 (32.6)</td><td>11 (32.4)</td><td>0.74 (0.22; 2.46)</td></tr><tr><td rowspan="3">PFNA</td><td>Tercile 1 (&lt;0.294)</td><td>19 (38.8)</td><td>9 (24.3)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.294-0.409])</td><td>12 (24.5)</td><td>20 (54.1)</td><td>3.13 (1.01; 10.43)</td></tr><tr><td>Tercile 3 (&ge;0.409)</td><td>18 (36.7)</td><td>8 (21.6)</td><td>0.65 (0.18; 2.22)</td></tr><tr><td rowspan="3">PFOA</td><td>Tercile 1 (&lt;1.808)</td><td>17 (33.3)</td><td>11 (29.7)</td><td>ref.</td></tr><tr><td>Tercile 2 ([1.808-2.557])</td><td>14 (27.5)</td><td>16 (43.2)</td><td>1.55 (0.51; 4.81)</td></tr><tr><td>Tercile 3 (&ge;2.557)</td><td>20 (39.2)</td><td>10 (27.0)</td><td>0.52 (0.15; 1.72)</td></tr><tr><td rowspan="3">PFUdA</td><td>Tercile 1 (&le;0.051)</td><td>17 (33.3)</td><td>10 (29.4)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.051-0.080])</td><td>20 (39.2)</td><td>13 (38.2)</td><td>0.97 (0.30; 3.11)</td></tr><tr><td>Tercile 3 (&gt;0.080)</td><td>14 (27.5)</td><td>11 (32.4)</td><td>1.25 (0.38; 4.16)</td></tr><tr><td rowspan="3">PFHxS</td><td>Tercile 1 (&lt;0.456)</td><td>18 (35.3)</td><td>10 (27.0)</td><td>ref.</td></tr><tr><td>Tercile 2 ([0.456-0.647])</td><td>13 (25.5)</td><td>16 (43.2)</td><td>1.80 (0.57; 5.80)</td></tr><tr><td>Tercile 3 (&ge;0.647)</td><td>20 (39.2)</td><td>11 (29.7)</td><td>0.71 (0.22; 2.21)</td></tr><tr><td rowspan="3">PFOS</td><td>Tercile 1 (&lt;3.736)</td><td>15 (29.4)</td><td>10 (27.0)</td><td>ref.</td></tr><tr><td>Tercile 2 ([3.736-5.074])</td><td>16 (31.4)</td><td>11 (29.7)</td><td>0.99 (0.29; 3.34)</td></tr><tr><td>Tercile 3 (&ge;5.074)</td><td>20 (39.2)</td><td>16 (43.2)</td><td>1.03 (0.33; 3.20)</td></tr><tr><td>PFAS mixture</td><td></td><td></td><td></td><td>1.33 (0.45; 4.07)</td></tr><tr><td>ΣPOP mixture</td><td></td><td></td><td></td><td>0.59 (0.30; 1.16)</td></tr><tr><td colspan="4"></td><td>0.48 (0.17; 1.38)</td></tr></tbody></table>

POP: persistent organic pollutant; Ref: reference; LOD: Limit Of Detection, p,p'-DDE: Dichlorodiphenyldichloroethylene, p,p'-DDT: Dichlorodiphenyltrichloroethane, HCH: Hexachlorocyclohexane, HCB: Hexachlorobenzene, PFBA: perfluorobutanoic acid, PFBS: Perfluorobutanesulfonic acid, PFDA: perfluorodecanoic acid, PFHxA: perfluorohexanoic acid, PFHxS: Perfluorohexanesulfonic acid, PFNA: perfluorononanoic acid, PFOA: perfluorooctanoic acid, PFOS: Perfluorooctanesulfonic acid, PFUdA: perfluoroundecanoic acid.

<sup>a</sup> Adjusted on maternal age at delivery, maternal pre-pregnancy body mass index, prenatal smoking status, prenatal maternal education level.

<sup>b</sup> OR are given for natural log-transformed POP analysed continuously when linearity was accepted.

<sup>c</sup> Effect for one quartile increase of all included chemicals using quantile g-computation.
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