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Response to Comments on “Endocrine Disrupting Nonylphenols Are Ubiquitous in Food” Alkylphenols are very relevant endocrine disrupting chemicals. They are degradation products of the industrially important alkylphenol ethoxylates (APE). APE are chemicals with outstanding surfactant properties. Ninety percent of the APE produced are nonylphenol ethoxylates (NPE). This means that the NPE are by far the most important group of the APE and also that the nonylphenols (NP) are the most significant group of APE metabolites. Because of their manufacturing process, the NP are a very complex mixture of isomers, and the composition can differ from manufacturer to manufacturer. The technical synthesis of NP starts from phenol, which is alkylated with a mixture of nonene isomers in an acid-catalyzed process. The chemical composition of technical nonene leads to a mixture of 4-NP consisting of isomeric compounds with differently branched structures of the nonyl side chain. In the toxicological studies cited by Degen and Bolt, only not characterized technical mixtures of nonylphenols (1, 2) or the much less relevant octylphenol (3, 4) have been investigated. This is, however, not adequate for precise toxicological assessment since the estrogenic effects of the individual nonylphenol isomerssas has been shown by our recent studiessis heavily dependent on the structure of the side chain. To conduct these investigations, a range of 25 different NP isomers were synthesized, and their estrogenic effects were determined using a well-known yeast assay (5). The assay uses a genetically modified yeast strain in which two plasmids have been inserted. The first generates the human R-estrogen receptor. When the test substance is added, it docks with different degrees of intensity to the receptor, and this then correspondingly modifies its structure. This receptor complex then binds with the second plasmid, depending on the extent to which the structure was modified. The second plasmid then controls the formation of a marker enzyme, the activity of which can be measured. The result is a measure of the enzymatic activity, which is expressed in Miller units in this test (6). The Miller units are approximately proportional to the estrogenic effect of the test substance added. This test revealed activities for the different isomers of between 0 Miller units for linear nonylphenol and up to 40 000 Miller units for a heavily branched isomer. In summary, therefore, it can be said that the estrogenic effects of NP are heavily dependent on the structure and that isomer-specific toxicological studies and analysis is absolutely important! For this reason, the toxicological evaluation and analysis of this class of substances has the character of a multicomponent problem.
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During the course of this basic NP research, we developed a practical numbering system for all possible isomers based on their IUPAC names (5). Theoretically, there are more than 200 constitutional isomers. This is further complicated by the fact that many of the constitutional isomers have chiral C-atoms (up to 3 per isomer) and that optical isomers also have to be borne in mind, which may also have different biological effects. All these factors mean that more than 500 compounds are possible. Of these, we estimate that approximately 50-80 NP are present at the same time in biological and other environmentally relevant matrixes. The chemical structure of some of these has still not yet been elucidated, and until now, only few examinations were enforced to the variability of isomer distribution in food and other matrixes (contamination by different technical products, isomer-dependent and enantiomer-dependent degradation). Thus, there is an urgent need in the future to do a great deal more research into this important substance class. Only then will it be possible to conduct the discussion of the NP problem on a modern scientific level. We can summarize the future topics in NP research as follows: (i) The complete elucidation of the chemical structures of all NP isomers found in food and the environment including the synthesis of selected important compounds (ii) Isomer- and enantiomer-specific determination of NP in food and elucidation of the variability of isomer distribution (iii) Exact toxicological in vivo and in vitro studies by using defined environmentally relevant nonylphenol isomers with known chemical structures
Literature Cited (1) Nagao, T.; Wada, K.; Marumo, H.; Yoshimura, S.; Ono, H. Reprod. Toxicol. 2001, 15, 293. (2) Nagao, T.; Saito, Y.; Usumi, K.; Nakagomi, M.; Yoshimura, S.; Ono, H. Hum. Exp. Toxicol. 2000, 19, 284. (3) Degen, G. H.; Janning, P.; Wittsiepe, J.; Upmeier, A.; Bolt, H. M. Toxicol. Lett. 2002, 127, 225. (4) Fisher, J. S.; Turner, K. J.; Brown, D.; Sharpe, R. M. Environ. Health Perspect. 1999, 107, 397. (5) Guenther, K.; et al., in preparation. (6) Breithofer, A.; Graumann, K.; Scicchitano, M. S.; Karathanasis, S. K.; Butt, T. R.; Jungbauer, A. J. Steroid Biochem. Mol. Biol. 1998, 67, 421.
Klaus Guenther,* Volkmar Heinke, Bjoern Thiele, Einhard Kleist, Hartmut Prast, and Torsten Raecker Institute of Chemistry and Dynamics of the Geosphere Institute III: Phytosphere Research Centre Juelich D-52425 Juelich, Germany ES0300057
10.1021/es0300057 CCC: $25.00
2003 American Chemical Society Published on Web 05/03/2003
