Ronald L. Joiner

Effects of p-nonylphenol (np) and diethylstilboestrol (des) on the alderley park (alpk) rat: comparison of mammary gland and uterus sensitivity following oral gavage or implanted mini-pumps

An earlier report by Colerangle and Roy indicated that administration of p‐nonylphenol (NP) to Noble rats, via subcutaneously implanted mini‐pumps at estimated doses of 53.2 and 0.073 mg kg−1 day−1 for 11 days, led to proliferation of the mammary gland. Those results indicated a ca. 600‐fold enhancement in assay sensitivity to NP over that of the standard 3‐day rat uterotrophic assay. The potential importance of these observations led us to repeat the experiments in the Noble rat, as described earlier. Although our earlier results confirmed the reported effects of diethylsti lboestrol (DES) on the mammary gland of Noble rats, we found no effects with NP. The present report extends our investigations of the effects of NP and DES on the mammary gland and uterus of other rat strains using both oral dosing and exposure via mini‐pumps.

Absorption, bioavailability, and metabolism of para-nonylphenol in the rat

To better interpret the responses to para-nonylphenol (NP; CASRN84852-15-3) in in vivo toxicity studies, including estrogen-like activity, the bioavailability of 14C-radiolabelled NP has been determined in male and female CD rats following either single oral doses of 10 and 100 mg/kg, single i.v. doses of 10 mg/kg, or repeated daily oral doses of 10 mg/kg for up to 14 d. Up to 80% of an oral dose of NP was rapidly absorbed, the remainder being excreted unchanged in faeces. Excretion was largely complete within 24 h of dosing. Following absorption, NP was metabolised in the liver, with the majority of the metabolites excreted in bile, mainly as glucuronide conjugates. Unchanged NP was found only in bile and urine from female rats given a 100 mg/kg dose, indicating that metabolic saturation occurred. Following repeated dosing, steady state was reached within 7 d. There was no evidence of significant accumulation into tissue compartments nor of a significant change in clearance or the metabolite profiles in urine. These data suggest that the estrogen-like effects observed in toxicity studies with female rats at oral NP doses of approximately 50 mg/kg/d and greater are a result of the increased bioavailability of NP which occurs following metabolic saturation.

Comparative in vitro percutaneous absorption of nonylphenol and nonylphenol ethoxylates (NPE-4 and NPE-9) through human, porcine and rat skin

The purpose of this study was to assess the percutaneous absorption of nonylphenol (NP) and the nonylphenol ethoxylates, NPE-4 and NPE-9, in human, porcine and rat skin. In vitro studies with the NPEs were conducted for 8 h in flowthrough diffusion cells using topical solutions of 0.1, 1.0 and 10% in PEG-400 or 1% in water (NPE-9 only). NP absorption was assessed as a 1% solution in PEG-400. All compounds were 14C ring-labeled and radioactivity in perfusate was monitored over time. Skin deposition was measured at the termination of the experiment. Absorption into perfusate and total penetration (compound absorbed plus compound sequestered in skin) were calculated. Absorption of NPE-4, NPE-9 and NP was similar across all species at less than 1% of the applied dose over 8 h. Penetration was generally below 5% of applied dose, the majority located in the stratum corneum. In all species and for both NPEs, the fraction of dose absorbed was highest for the lowest applied dose. Absorptions expressed as actual mass absorbed over 8 h were similar (approximately 0.3 μg/cm2) across all concentrations. Penetration, but not absorption, was greater from a water vehicle compared to a PEG-400 vehicle, particularly in rat skin. These studies suggest that NP, NPE-4 and NPE-9 were minimally absorbed across skin from all three species. Fractional absorption was concentration-dependent, making the actual absorbed flux constant across all doses.

One-generation reproductive toxicity study of dietary 17β-estradiol (E2; CAS No. 50-28-2) in CD-1® (Swiss) mice

There is no information on reproductive/developmental effects in mice from dietary estrogen. Therefore, 10 adult CD-1 mice/sex/group were administered dietary 17beta-estradiol (E2) at 0, 0.005, 0.05, 0.5, 2.5, 5, 10, and 50 ppm for 2-week prebreed, mating, gestation, lactation. F1 weanlings (3/sex/litter) were necropsied and 2/sex/litter were retained, with exposure, until vaginal patency (VP) or preputial separation (PPS) and then necropsied. Results included complete infertility at 2.5-50 ppm with normal mating indices. At 0.5 ppm (and above), F0 adult female uterus plus cervix plus vagina weights (UCVW) were increased. At 0.5 ppm: prolonged gestational length; increased F1 stillbirth index; reduced live birth index and litter size; decreased testes and epididymides weights at weaning; unaffected AGD on pnd 0 and 21; delayed PPS; increased undescended testes; unaffected prostate weight; accelerated VP; enlarged vaginas; fluid-filled uteri. At 0.05 ppm: no F0 reproductive effects, increased F1 weanling UCVW; delayed PPS. The NOEL was 0.005 ppm ( approximately 1 microg/kg/day).

The uterotrophic activity of nonylphenol in the rat is not mediated by aromatase enzyme induction

p-Nonylphenol (NP) is weakly estrogenic to rodents and to some species of fish. All evidence to date has indicated that the estrogenic effects of NP are due to the interaction of NP with the estrogen receptor. Recent findings of increased plasma estradiol in fish exposed to NP have, however, led to the proposal of an alternative mechanism for NP-induced estrogenicity in this species, possibly via induction of aromatase enzymes. In the present studies, this hypothesis was investigated in rats using the aromatase inhibitor anastrozole. The results indicated that the uterotrophic action of NP, as with estradiol used as a positive control, is mediated directly by its interaction with uterine ER, rather than an indirect effect via aromatase enzyme induction. Circulating levels of estradiol were unchanged after NP treatment and the aromatase inhibitor anastrozole failed to inhibit NP-induced uterine growth. These results are consistent with previous published data on NP in rodents.

In Vitro Percutaneous Absorption of Nonylphenol (NP) and Nonylphenol Ethoxylates (NPE-4 and NPE-9) in Isolated Perfused Skin

Skin contact with nonylphenol ethoxylates (NPE), a group of widely used surfactants, is the primary source of human exposure. Previous studies have shown that the absorption of NPE through human and animal skin in vitro is limited (<1% over 8 hr) [Monteiro-Riviere et al. Toxicol Indust Health 2000; 16:49–57]. The purpose of this study was to examine the percutaneous absorption of NPE and the chemical precursor, nonylphenol (NP), in the isolated perfused porcine skin flap (IPPSF) model for comparison to the in vitro porcine skin flow through (PSFT) diffusion studies. The IPPSF model is considered to accurately predict absorption of chemicals through human skin. The IPPSF was dosed with 100 μl of 1% 14C ring-labeled NP, 14C ring-labeled NPE-4, or 14C ring-labeled NPE-9 in aqueous polyethylene glycol (PEG-400) solution and perfused for 8 hr. All three chemicals were minimally absorbed, with only approximately 0.1% of the applied dose found in the perfusate over the 8-hr collection. This absorbed material represents the systemic exposure expected following skin contact in humans. In addition, less than 1% of the applied dose penetrated into the stratum corneum and underlying dermis, but remained within the skin and did not go through to the perfusate. Thus, the overall potential systemic exposure to these chemicals from skin contact, using a model considered similar to human skin in vivo, is less than 1%. The absorption results of this study were consistent with previous studies in the PSFT model. The penetration of NPEs and NP in the IPPSF was less than the PSFT and is probably more predictive of in vivo human absorption as this model is physiologically closer to human skin. This suggests that the overall potential for skin absorption of these chemicals in humans is even lower than previous estimates.