Evaluating Reproductive Impairment Induced by Phthalates: A Systematic Preclinical Meta-Analysis of Wistar Rat Models with A Qualitative Assessment of Human Regulatory Frameworks

Abstract

Phthalates represent a pervasive class of endocrine-disrupting chemicals found throughout the biosphere. While the Wistar rat is the widely used animal model for assessing phthalate-related reproductive toxicity, a cohesive synthesis of these preclinical data to inform hazard identification is notably absent. This study aimed to systematically meta-analyze the reproductive impacts of phthalates in Wistar rats, qualitatively correlate these findings with current regulatory safety thresholds as a means of identifying research gaps, and assess their consistency with human epidemiological observations. Adhering to Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) protocols, we conducted an exhaustive search of PubMed, Web of Science, Scopus, and TOXLINE for primary research published between 2000 and 2025. Statistical analysis utilized random-effects models to calculate standardized mean differences (SMDs), while study rigor was evaluated using the SYRCLE Risk of Bias tool and the Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach. Out of 1,847 initial records, 82 studies met the inclusion criteria. Our analysis revealed that perinatal exposure to di-(2-ethylhexyl) phthalate (DEHP) and dibutyl phthalate (DBP) resulted in a statistically significant reduction in anogenital distance among male offspring (SMD: −1.24; 95% CI: −1.58 to −0.90; moderate certainty). Furthermore, sperm motility exhibited a sharp decline (SMD: −0.98; 95% CI: −1.24 to −0.72; high certainty) within just twenty-one days of exposure. Histopathological assessments consistently identified Leydig cell abnormalities, seminiferous tubule degradation, and elevated oxidative stress markers. Notably, qualitative benchmarking of these findings against U.S. Environmental Protection Agency (EPA) reference doses indicated that adverse reproductive outcomes occurred at exposure levels that appear lower than existing regulatory benchmarks when compared directly, though formal interspecies extrapolation (allometric scaling, toxicokinetic differences, and uncertainty factors) is required before drawing definitive conclusions about human risk. Molecular docking simulations corroborated these outcomes, demonstrating clear DEHP-androgen receptor binding, thereby supporting a potential mechanism of anti-androgenic activity. These findings illuminate critical areas for further investigation in regulatory science. We propose that integrating systematic preclinical synthesis into modern risk assessment frameworks is essential to improving the accuracy and protective capacity of public health policies regarding phthalate exposure, while acknowledging that animal-to-human extrapolation remains a significant challenge. Registration: PROSPERO (CRD420261358797).

HIGHLIGHTS

• A systematic synthesis of 82 Wistar-rat studies identified reproducible phthalate-related reproductive effects.
• Perinatal exposure was associated with reduced anogenital distance and increased nipple retention in male offspring.
• Phthalate exposure was associated with reduced sperm motility, sperm count, testis weight, and reproductive success.
• Oxidative stress, Leydig-cell dysfunction, and androgen-receptor interactions provide convergent mechanistic evidence.
• Qualitative comparison with regulatory benchmarks highlights the need for formal interspecies extrapolation in risk assessment.

GRAPHICAL ABSTRACT