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Key value for chemical safety assessment

Effects on fertility

Description of key information

Experimental animals  

A three generation study performed in mice (strain not specified) administered with 250 mg/kg test item in chocolate revealed no drug related teratogenic or reproductive deficits [1]. The estrogenic activity of the test substance was studied in immature Wistar rats. Estrogenic response in terms of increase in the glycogen content in the uterus was obtained at 4 mg test item. The minimum effective concentration of diethylstilbestrol was about 0.1 mg in under the conditions used in this assay [2]. Subcutaneous injection of the test item stimulates the growth of the immature rat uterus as shown in a similar study [3]. Thirteen-week toxicity studies of the test substance were performed using F344/N rats and B6C3F1 mice. Rats and mice were fed ad libitum with a NIH 07 diet containing 0; 3000; 6000; 12,000; 25,000; or 50,000 ppm test item. The primary treatment-related findings that occurred during the mouse studies involved the reproductive and hematopoietic systems. Reproductive changes including depressed testis and right epididymal weights and sperm density, an elevated production of abnormal sperm, and morphologic alterations in seminiferous tubules occurred at all levels of exposure (3000-50,000 ppm) [4]. In a 27-week study in mice the data obtained show atrophy of the seminiferous tubules in the testis, hyperplasia of the testicular interstitial (Leydig) cells, and epididymal hypospermia occurred in most males dosed at 3,000 and 12,000 ppm in feed [5]. The potential reproductive toxicity of the test substance was evaluated using Swiss CD-1 mice in the RACB protocol because of literature suggestions that the test item acts as a weak oestrogen, and because data gathered at the end of a 90-day NTP study in B6C3F1 mice indicated adverse male effects [5]. Concentrations of the test item in feed for this study were 0.1, 0.7, and 3.0 % weight per volume. In summary, the test substance at these concentrations produced significant reproductive toxicity (fewer litters per pair, and fewer pups per litter) in the absence of changes in body or somatic organ weights. The second generation did not appear more affected than the first [6]. 

Human data  

The test substance competed with oestrogen binding affinity in MCF-7 human breast cancer cells with a binding affinity of 10-4 that of oestradiol and stimulated cell growth in vitro. The estrogenic activity leads to elevated progesterone receptor levels in the MCF-7 cells. The anti-oestrogen 4-hydroxytamoxifen blocked the growth stimulation triggered by the test substance [7]. To the best of our current knowledge, no further information on reproductive or developmental related toxicity of the test substance in humans has been reported. 


1.    Stockinger, L., [Experiments on the analysis of the effects of phenolphthalein]. Arzneimittelforschung, 1965. 15(5): p. 550-4.

2.    Bitman, J. and H.C. Cecil, Estrogenic activity of DDT analogs and polychlorinated biphenyls. J Agric Food Chem, 1970. 18(6): p. 1108-12.

3.    Nieto, A., C. Garcia, and M.S. Lopez de Haro, In vivo estrogenic and antiestrogenic activity of phenolphthalein and derivative compounds. Biochem Int, 1990. 21(2): p. 305-11.

4.    Dietz, D.D., et al., Subchronic (13-week) toxicity studies of oral phenolphthalein in Fischer 344 rats and B6C3F1 mice. Fundam Appl Toxicol, 1992. 18(1): p. 48-58.

5.    National Toxicology Program (NTP), Toxicology and Carcinogenesis Studies of Phenolphthalein (CAS no. 77-09-8) in F344/N Rats and B6C3F1 Mice (Feed Studies) - Technical Report 465, in Technical Report 465. 1996.

6.    Reproductive toxicology. Phenolphthalein. Environ Health Perspect, 1997. 105 Suppl 1: p. 335-6.

7.    Ravdin, P.M., M. van Beurden, and V.C. Jordan, Estrogenic effects of phenolphthalein on human breast cancer cells in vitro. Breast Cancer Res Treat, 1987. 9(2): p. 151-4. 

Justification for classification or non-classification

Classification, Labeling, and Packaging Regulation (EC) No. 1272/2008

The substance is harmonized classified for reproductive toxicity Cat.2 (H361f) according to Annex VI of Regulation (EC) No 1272/2008 (CLP Regulation).

Additional information