Since bisphenol F (4,4'-dihydroxydiphenylmethane) has been reported to exhibit estrogen agonistic properties in the uterotrophic assay, we performed a 28-day repeated-dose toxicity study (enhanced OECD test guideline No. 407) on bisphenol F based on the OECD draft protocols to determine whether it has endocrine-mediated properties. Bisphenol F was orally administered at doses 0, 20, 100 and 500 mg/kg per day for at least 28 days, but no clear endocrine-mediated changes were detected, and it was concluded to have no endocrine-mediated effects in young adult rats. On the other hand, the main effect of bisphenol F was concluded to be liver toxicity based on clinical biochemical parameters and liver weight, but without histopathological changes. The no-observed-effect level for bisphenol F is concluded to be under 20 mg/kg per day since decreased body weight accompanied by decreased serum total cholesterol, glucose, and albumin values were observed in the female rats given 20 mg/kg per day or higher doses of bisphenol F.

The murine local lymph node assay (LLNA) is currently recognized as a stand-alone sensitization test for determining the sensitizing potential of chemicals, and it has the advantage of yielding a quantitative endpoint that can be used to predict the sensitization potency of chemicals. The EC3 has been proposed as a parameter for classifying chemicals according to the sensitization potency. We previously developed a non-radioisotopic endpoint for the LLNA based on 5-bromo-2'-deoxyuridine (BrdU) incorporation (non-RI LLNA), and we are proposing a new procedure to predict the sensitization potency of chemicals based on comparisons with known human contact allergens. Nine chemicals (i.e. diphencyclopropenone, p-phenylenediamine, glutaraldehyde, cinnamicaldehyde, citral, eugenol, isopropyl myristate, propyleneglycol and hexane) categorized as human contact allergen classes 1-5 were tested by the non-RI LLNA with the following reference allergens: 2,4-dinitrochlorobenzene (DNCB) as a class 1 human contact allergen, isoeugenol as a class 2 human contact allergen and alpha-hexylcinnamic aldehyde (HCA) as a class 3 human contact allergen. Consequently, nine test chemicals were almost assigned to their correct allergen class. The results suggested that the new procedure for non-RI LLNA can provide correct sensitization potency data. Sensitization potency data are useful for evaluating the sensitization risk to humans of exposure to new chemical products. Accordingly, this approach would be an effective modification of LLNA with regard to its experimental design. Moreover, this procedure can be applied also to the standard LLNA with radioisotopes and to other modifications of the LLNA.

A two-generation reproductive toxicity study with extra parameters was performed for Butyl benzyl phthalate (BBP). The compound was administered orally by gavage with the doses of 0, 100, 200, or 400 mg/kg/day to groups of 24 Crj:CD (SD)IGS rats of both sexes to confirm the utility of the protocol for identification of non-steroid chemicals with endocrine activity by ssessing effects on parental animals and offspring. Softening of the testes, diffuse atrophy of testicular seminiferous tubules, decreased spermatozoa and/or residual germ cells in the epididymal lumina were observed in the F1generation after doses more than 100 mg/kg, lowering of the F1 epididymal weights at doses more than 200 mg/kg, along with low F0 epididymal weights, Leydig cell hyperplasia, residual germ cells in the epidimymal lumina, and low seminal vesicle weights, small testes and epididymes, partial aplasia or aplasia of the epididymes, and Leydig cell hyperplasia in the F1 generation with 400 mg/kg. With regard to effects on the reproductive capacity, F1 parents at the dose of 400mg/kg showed a reduced fertility index and delayed preputial separation of the penis. In the offspring, lowered body weights in the F1 case, and change in anogenital distance in the F1 females and F2 males were observed at doses more than 100 mg/kg, with low splenic weights at 400 mg/kg in both generations. Thus, the utility of this protocol was confirmed. In the parental animals, the no observed effect level (NOEL) and the no observed adverse effect level (NOAEL) were less than 100 mg/kg/day, and no serious effects on the reproductive capacity were induced at doses less than 200 mg/kg/day. The NOEL and NOAEL for the growth and development of offspring were concluded to be less than 100 mg/kg/day.

We performed a 28-day repeated-dose toxicity study of diethylphthalate based on the draft protocol of the "Enhanced OECD Test Guideline 407" to investigate whether it has endocrine-mediated properties according to this assay. Diethylphthalate was orally administered to SD rats at doses of 0, 40, 200, and 1,000 mg/kg/day for at least 28 days, but no endocrine-mediated effects were detected based on any of the parameters examined, suggesting that diethylphthalate does not possess endocrine properties according to this assay.

We performed a 28-day repeated-dose toxicity study of di(2-ethylhexyl)adipate (DEHA) based on the draft protocol of the "Enhanced OECD Test Guideline 407" to investigate whether it has endocrine-mediated properties according to this assay. DEHA was orally administered to SD rats at doses of 0, 40, 200 and 1,000 mg/kg/day for at least 28 days, and disturbance of the estrous cycle and increased ovarian follicle atresia were detected in the 1,000 mg/kg group.