Quantification of per- and polyfluoroalkyl substances in plasma and follicular fluid of patients undergoing in vitro fertilization in Iowa: A pilot study.

This retrospective cohort study was conducted with use of previously collected plasma and follicular fluid from the University of Iowa tissue bank (Institutional Review Board #201709794) from patients with an infertility diagnosis of polycystic ovary syndrome (PCOS), endometriosis, or unexplained infertility undergoing IVF between September 2019 and September 2023. Demographic data included age, body mass index, race, gravidity, and parity. Fertility outcomes included insemination method, fertilization rate, total metaphase II oocytes, total blastocysts meeting transfer or cryopreservation criteria, and cumulative live birth rate. Using solid-phase extraction and liquid chromatography-tandem mass spectrometry, we surveyed plasma and follicular fluid for 43 PFAS, including legacy and replacement species (Supplemental Tables S1–S3). Statistical analysis was performed with SPSS v29. Quantitative data are presented as means (± SD) and categorical data as frequencies. For each analyte, we report the percentage of the samples found to have the analyte above the limit of quantification (LOQ) and the concentration of the analyte among those samples with a concentration above the LOQ. Normality of continuous data was assessed through review of histograms and comparison of 5% trimmed mean to the mean. Homoscedasticity, or equal variance across groups, was assessed with Levene’s test. ANOVA was used to compare continuous data when assumptions of normality and homoscedasticity were met. Welch’s ANOVA was used when the assumption of homoscedasticity was violated. The Χ 2 -square test was used to evaluate differences in categorical factors. Tukey’s HSD test was applied between infertility groups.

Prapti Singh-Herren: Writing – review & editing, Writing – original draft, Visualization, Validation, Resources, Project administration, Methodology, Investigation, Conceptualization. Samantha Good: Writing – review & editing, Visualization, Validation, Methodology, Investigation, Formal analysis, Data curation. Karen M. Summers: Writing – review & editing, Visualization, Validation, Software, Resources, Project administration, Methodology, Investigation, Formal analysis, Data curation. Joseph A. Charbonnet: Writing – review & editing, Validation, Methodology, Investigation. Amy E. Sparks: Writing – review & editing, Methodology, Conceptualization. Aileen F. Keating: Writing – review & editing, Validation, Supervision, Resources, Methodology, Conceptualization.

This study had 14 subjects in each diagnosis group: PCOS (A), endometriosis (B), and unexplained infertility (C). Mean age was similar between groups. Body mass index was highest in the PCOS group, but did not differ among groups after Tukey post-hoc comparison (A, 29.74 ± 5.86; B, 25.30 ± 3.25; C, 25.17 ± 6.26 kg/m 2 ; P =.04; Tukey post-hoc A:B P =.08, A:C P =.07 and B:C P =.998). Median gravidity was 0 with an interquartile range (IQR) of (0, 1) for gravidity in the PCOS group, and an IQR of (0, 0) in both other groups ( P =.262). Median parity and IQR was 0 (0, 0) for all groups. Most patients (86%) identified as White and had no prior live birth. Cumulative live birth rate was similar among groups (A, 93%; B, 93%; C, 86%; Table 1 ). Of the 43 PFAS targeted in these patients, 21 species in plasma and 23 species in follicular fluid were quantified ( Fig. 1 A,B). Twelve species were quantified among ≥ 50% of patients [6:2 FTS, PFBS, and PFMBA (plasma only); PFBA and PFPeS (follicular fluid only); and linear and branched (L + br) PFHxS, L + br PFOS, PFDA, PFHpA, PFHpS, PFHxA, PFNA, PFOA, and PFUdA (both matrices)]. For each of these 12 PFAS, there were no differences in plasma or follicular fluid concentrations among the infertility diagnosis groups ( P values, .18–.94). Five species were detected in 100% of plasma samples (L+br PFOS, PFOA, L+br PFHxS, PFNA, and PFHpS), and three species were detected in 100% of follicularfluid samples (L+br PFOS, PFOA, and L+br PFHxS). PFOA and L + br PFHxS were present in all patients. PFAS concentrations were consistently higher in plasma than follicular fluid within the same patient. Table 1 Patient Characteristics PCOS ( n =14) Endometriosis ( n =14) Unexplained infertility ( n =14) P value Age 30 ± 2.32 30 ± 2.31 31 ± 2.06 .644 BMI 29.74 ± 5.86 25.30 ± 3.25 25.17 ± 6.26 .044 Duration of infertility 3.36 ± 1.78 3.21 ± 1.85 3.64 ± 2.21 .840 Comorbidities a 5 (36%) 2 (14%) 5 (36%) ENM Gravidity 0.43 ± 0.76 0.07 ± 0.27 0.14 ± 0.36 .270 b Parity 0.07 ± 0.27 0 ± 0 0.07 ± 0.27 .610 Race ENM  White 12 (86%) 12 (86%) 12 (86%)  Black 1 (7%) 0 (0%) 0 (0%)  Hispanic 0 (0%) 1 (7%) 0 (0%)  Asian 1 (7%) 1 (7%) 2 (14%) # of oocytes retrieved 27 ± 13.15 17 ± 13.99 22 ± 15.40 0.21 Insemination method ENM  Standard 8 (57%) 7 (50%) 7 (50%)  ICSI 5 (36%) 7 (50%) 7 (50%)  Both 1 (7%) 0 (0%) 0 (0%) Fertilization rate 65.3 ± 18.7 69.1 ± 16.0 71.1 ± 23.4 0.73 Mature oocytes (M2) 15 ± 6.20 7 ± 9.16 13 ± 10.44 0.05 Total blastocysts cryopreserved or transferred 8 ± 5.16 5 ± 4.18 7 ± 5.08 0.52 Cumulative live birth rate 13 (93%) 13 (93%) 12 (86%) ENM Cumulative clinical pregnancy rate c 87.5 ±25.5 74.4 ± 37.1 59.8 ± 30.8 .078 Cumulative implantation rate d 87.5 ± 43.0 77.4 ± 37.8 57.4 ± 31.5 .112 ENM, Expectations of statistical test not met. a Comorbidities include history of cervical cancer, ankylosing spondylitis, Asherman syndrome, diabetes, history of recurrent cyst removal, hypothyroidism, lyme disease, thyroid nodules. b p value for Welch test. c Defined at participant level as number of clinical pregnancies/embryo transfer cycles. d Defined at participant level as number of fetal heartbeats on ultrasound/embryos transferred (both across all cycles). Figure 1 PFAS concentrations. ( A ) Plasma PFAS. Plasma PFAS concentrations (ng/L) > LOQ quantified in > 50% of samples. Boxplots represent the interquartile range (IQR), with whiskers extending to 1.5 × IQR and the middle line indicating the median. Circles represent individual measurements, with circle and boxplot color indicating the percentage of samples in which each analyte was quantified. L+br indicates sum linear and branched isomers. ( B ) Follicular fluid PFAS. Follicular fluid PFAS concentrations (ng/L) > LOQ quantified in > 50% of samples. Boxplots represent the interquartile range (IQR), with whiskers extending to 1.5 × IQR and the middle line indicating the median. Circles represent individual measurements, with circle and boxplot color indicating the percentage of samples in which each analyte was quantified. L+br indicates sum linear and branched isomers. Patient Characteristics ENM, Expectations of statistical test not met. Comorbidities include history of cervical cancer, ankylosing spondylitis, Asherman syndrome, diabetes, history of recurrent cyst removal, hypothyroidism, lyme disease, thyroid nodules. p value for Welch test. Defined at participant level as number of clinical pregnancies/embryo transfer cycles. Defined at participant level as number of fetal heartbeats on ultrasound/embryos transferred (both across all cycles). PFAS concentrations. ( A ) Plasma PFAS. Plasma PFAS concentrations (ng/L) > LOQ quantified in > 50% of samples. Boxplots represent the interquartile range (IQR), with whiskers extending to 1.5 × IQR and the middle line indicating the median. Circles represent individual measurements, with circle and boxplot color indicating the percentage of samples in which each analyte was quantified. L+br indicates sum linear and branched isomers. ( B ) Follicular fluid PFAS. Follicular fluid PFAS concentrations (ng/L) > LOQ quantified in > 50% of samples. Boxplots represent the interquartile range (IQR), with whiskers extending to 1.5 × IQR and the middle line indicating the median. Circles represent individual measurements, with circle and boxplot color indicating the percentage of samples in which each analyte was quantified. L+br indicates sum linear and branched isomers.

Our pilot study demonstrates PFAS are quantifiable in plasma and follicular fluid of patients undergoing IVF from an academic fertility center in Iowa. This finding is consistent with previous international studies indicating the presence of PFAS in women pursuing fertility care ( 2 , 3 , 4 ). The endometriosis group had comparatively fewer total oocytes (17 ± 13.99, P =.21) and mature oocytes (7 ± 9.16, P =.05) compared with the other groups, although not statistically significant, this could be related to the inflammatory nature related to the pathophysiology of endometriosis. This finding was not recorded in a prior study which analyzed PFAS in endometriosis patients (1), yet decreased oocyte yield as an outcome related to endometriosis itself has been noted, providing evidence that our data are in agreement with other findings. No significant differences were present in the most common PFAS among the three groups. This also was true for the treatment outcomes among groups, yet we were not powered to assess these outcomes. Strengths of this study include the first report in our region of PFAS in this patient population among three common diagnosis groups. Demographics were similar among diagnosis groups. Limitations include the difficulty of identifying a fitting control group and the unique pathophysiology of the included diagnoses being a potential confounding factor, thus limiting the interpretation of our results. Unlike a previous study related to PCOS diagnoses ( 5 ), we did not determine association between an infertility diagnosis and the presence of a specific PFAS. The prevalence of PFAS among infertility patients pursuing IVF suggests further studies of these reproductive toxicants are needed to improve understanding their effects on both general and reproductive health.

P.S. has a professional development and medical advisory role with Vitrolife. S.G. has nothing to disclose. K.M.S. has nothing to disclose. J.A.C. has nothing to disclose. A.E.S. has nothing to disclose. A.F.K. has nothing to disclose.