Male preputial separation, failed/delayed

This state is characterized by failure or delay in the natural detachment of the prepuce from the glans penis.

The biological compartment this is measured in is the glans penis, glans-prepuce adhesion interface, and preputial epithelium.

Normal preputial separation serves protective, mechanical, immunological and erogenous functions (Paraboschi and Garriboli 2020).  Biological consequences of a failure in this separation fall into the same areas.   

Gross anatomical observations are a direct, standardized, and reproducible strategy and can be conducted periodically.  Binary scoring and ordinal scales can be employed to describe degree of separation and timing.

Taxonomic Applicability

Delayed/failed preputial separation is limited to mammals because the process is unique to mammalian foreskin anatomy.  It is well documented in rats and mice as a developmental milestone (Gray et al., 2001).  It also occurs in humans but isn’t understood as a standardized developmental marker (Cunha et al., 2020).

Lifestage Applicability

Preputial separation is a maturation event that takes place postnatally.  It is an indicator of the onset of puberty in rodents, typically occuring 4-5 weeks after birth in mice and 6-7 weeks after birth in rats.  In humans, it is a variable and ongoing process starting in infancy with significant physiological shifts coinciding with puberty.  In all cases, if separation has not occurred by adulthood, it has failed permanently.

Sex Applicability

Male preputial separation refers to the detachment of the foreskin from glans penis thereby limiting the applicability to the male.

A delay or failure in preputial separation represents an apical endpoint in standard developmental and reproductive toxicity tests which are used for hazard identification, risk assessment, and regulatory decision-making, especially as it relates to chemicals with potential endocrine-disrupting activity.

International Regulatory Context

PPS is included in several OECD Test Guidelines that are adopted by OECD member countries as well as several non-OECD countries that adhere to Mutual Acceptance of Data (MAD) e.g., Argentina, Brazil, India).

• OECD Test No. 443 - Extended One-Generation Reproductive Toxicity Study (EOGRTS)
• OECD Test No. 416 - Two-Generation Reproductive Toxicity Study:
  • Requires daily evaluation of balano-preputial separation in male pups.  It is used for comprehensive assessment of developmental and reproductive effects inclusive of endocrine disruption indicators.

OECD member countries and regions adhering to MAD use data on delayed PPS as a regulatory endpoint used for:

• • Hazard identification and classification (e.g., under EU CLP No 1272/2008)
  • Risk assessment supporting chemical registration/authorization decisions under REACH/ECHA.

United States Regulatory Context

• U.S. EPA Two-Generation Reproductive Toxicity Test Guideline (OPPTS 870.3800)
  • PPS is an endpoint used for evaluation under Federal Insecticide, Fungicide, and Rodenticide Act (FIFRA) and Toxic Substances Control Act (TSCA) to detect effects on the integrity and performance of the reproductive system.
• EPA Endocrine Disruptor Screening Program (EDSP) Tier 1 and Tier 2
  • Incorporates rodent pubertal assays (including male pubertal assay) that use age at preputial separation as a sensitive measure of disruption of androgen signaling.
    • EDSP is integrated into the Federal Food, Drug, and Cosmetic Act (FFDCA) in section 408(p) and the Safe Drinking Water Act in Section 1457.

EPA uses these tests to screen, identify, and prioritize chemicals for potential endocrine activity and hazard.  Regulatory decisions like restrictions, registrations, and risk mitigation occur under TSCA and FIFRA statutes.

Cunha, G. R., Sinclair, A., Cao, M., & Baskin, L. S. (2020). Development of the human prepuce and its innervation. Differentiation, 111, 22-40.

Federal Insecticide, Fungicide, and Rodenticide Act, 7 U.S.C. §§ 136–136y (2023).

Gray Jr, L. E., Ostby, J., Furr, J., Wolf, C. J., Lambright, C., Parks, L., ... & Guillette, L. (2001). Effects of environmental antiandrogens on reproductive development in experimental animals. Apmis, 109(S103), S302-S319.

OECD (2001), Test No. 416: Two-Generation Reproduction Toxicity, OECD Guidelines for the Testing of Chemicals, Section 4, OECD Publishing, Paris, https://doi.org/10.1787/9789264070868-en.

OECD (2025), Test No. 443: Extended One-Generation Reproductive Toxicity Study, OECD Guidelines for the Testing of Chemicals, Section 4, OECD Publishing, Paris, https://doi.org/10.1787/9789264185371-en.

Paraboschi, I., Garriboli, M. (2020). Medical and Surgical Uses of the Prepuce. In: Normal and Abnormal Prepuce. Springer, Cham. https://doi.org/10.1007/978-3-030-37621-5_8

Toxic Substances Control Act, as amended by the Frank R. Lautenberg Chemical Safety for the 21st Century Act, 15 U.S.C. §§ 2601–2697 (2023

U.S. Environmental Protection Agency. (2013, June 14). Endocrine disruptor screening program; final policies and procedures for screening Safe Drinking Water Act substances. Federal Register. https://www.federalregister.gov/documents/2013/06/14/2013-14228/endocrine-disruptor-screening-program-final-policies-and-procedures-for-screening-safe-drinking

U.S. Environmental Protection Agency. Final List of Initial Pesticide Active Ingredients and Pesticide Inert Ingredients to be Screened Under the Federal Food, Drug, and Cosmetic Act [Document ID EPA-HQ-OPPT-2004-0109-0080]. Regulations.gov. https://www.regulations.gov/document/EPA-HQ-OPPT-2004-0109-0080

U.S. Environmental Protection Agency, Office of Prevention, Pesticides and Toxic Substances. (1998). Health Effects Test Guidelines: OPPTS 870.3800—Reproduction and fertility effects. https://www.epa.gov/test-guidelines-pesticide-registration/series-870-health-effects-test-guidelines