The 2-phenylbenzotriazole-type water pollutant PBTA-2 has cytochalasin B-mimetic activity

The 2-phenylbenzotriazole (PBTA)-type water pollutant, 2-[2-(acetylamino)-4 -[N-(2-cyanoethyl)ethylamino]-5-methoxyphenyl]-5-amino-7-bromo-4-chloro-2H- benzotriazole (PBTA-2), has been recently identified in samples from the Ni shitakase River in Kyoto, Japan, and shows potent mutagenic activities in S almonella typhimurium in the presence of a microsomal metabolizing system ( S9 mix). In the present study, we conducted the in vitro micronucleus (MN) test on PBTA-2 in the absence and presence of S9 mix in two Chinese hamster cell Lines, CHL and V79-MZ. In the MN test, PBTA-2 was weakly positive in CHL cells and strongly positive in V79-MZ cells. Because the positive resul ts were accompanied by a statistically significant increase in the number o f polynuclear (PN) and/or mitotic (M) cells, we examined treated cells in m etaphase to see if numerical chromosome aberrations were being induced. We found that PBTA-2 induces polyploidy in both CHL and V79-MZ cells. A detail ed analysis of MN preparations showed that in CHL cells, PBTA-2 predominant ly induces equal-sized binucleated cells. Rhodamine phalloidin staining rev ealed that PBTA-2 causes actin filament abnormalities in both cell lines si milar to those caused by cytochalasin B. Cytochalasin B induced PN cells pr edominantly and dose dependently, and almost all the cells were equal-sized and binucleate. The results suggest that PBTA-2 has cytochalasin B-mimetic activity, although agents affecting actin filaments, such as cytochalasins , phallotoxins and chloropeptide, have been derived only from molds so far. This study also suggests that our MN test protocol may be used to identify chemicals that have cytochalasin B-mimetic activity as well as those that induce numerical aberrations. (C) 2000 Elsevier Science B.V. All rights res erved.