Safety evaluation of certain food additives - ipcs inchem

432 ALKOXY-SUBSTITUTED ALLYLBENZENES
of the strains tested, but not unequivocally positive (Ichinotsubo et al., 1981b; Venitt
& Crofton-Sleigh, 1981). In the SOS induction assay using E. coli PQ37, uniformly
negative results were obtained with safrole (Quillardet et al., 1985; Kevekordes
et al., 1999).
Equivocal results for safrole were obtained in the Bacillus subtilis rec assay.
At concentrations up to 20 000 μg/disc, no induction of recombination was observed
(Sekizawa & Shibamoto, 1982; Suter & Jaeger, 1982). In other studies in B.
subtilis, induction of recombination was reported at concentrations of 21 000 μg/
disc and greater (Kada et al., 1980; Kada, 1981).
Equivocal results were obtained in assays with various strains of S.
cerevisiae, including D3, D5, D6, D7, D61.M, JD1, PV-3, RS9, RS112, T1, T2, C658-
K42 and XV185-14C, to assess the induction of mitotic aneuploidy (Simmon, 1979b;
Jagannath et al., 1981; Kassinova et al., 1981; Parry & Sharp, 1981; Sharp & Parry,
1981b; Zimmermann & Scheel, 1981; To et al., 1982a; Arni, 1985; Brooks et al.,
1985; Ferguson, 1985; Inge-Vechtomov et al., 1985; Parry & Eckardt, 1985;
Zimmermann et al., 1985; Morita et al., 1989; Schiestl, 1989; Schiestl et al., 1989a,
1989b). Safrole was highly toxic to various strains of S. cerevisiae at concentrations
as low as 30 μg/ml (Zimmermann & Scheel, 1981; Arni, 1985; Zimmermann et al.,
1985; Schiestl et al., 1989a, 1989b). Positive results were often observed in studies
with modified protocols that used cytotoxic levels of safrole or that used nonstandard
strains of S. cerevisiae (Kassinova et al., 1981; Mehta & von Borstel, 1981;
Parry & Sharp, 1981; Sharp & Parry, 1981a, 1981b; Ferguson, 1985; Schiestl, 1989;
Schiestl et al., 1989a, 1989b).
Forward mutation assays of safrole in L5178Y mouse lymphoma cells were
predominantly negative (To et al., 1982a; Oberly et al., 1984, 1985; Lee & Webber,
1985; Myhr et al., 1985; Mitchell et al., 1988; Myhr & Caspary, 1988). Positive results
were often observed in the presence of metabolic activation (Jotz & Mitchell, 1981;
Garner & Campbell, 1985; Myhr et al., 1985; Styles et al., 1985; Mitchell et al., 1988;
Myhr & Caspary, 1988). In several of the assays where induction of forward
mutations was observed, cytotoxicity was also seen, and the mutation rate was less
than 2 times that of negative controls (Jotz & Mitchell, 1981; Myhr & Caspary, 1988).
One study reported inconclusive results based on the observations that the mutation
rate in the background was unusually high and that cytotoxicity was observed at
higher concentrations of safrole (Knaap & Langebroek, 1985).
Estragole at concentrations of 10 3 –10 5 mol/l did not induce the increase of
chromosomal aberrations in V79 cells when tested alone or with metabolic
activation or in primary rat hepatocytes (Müller et al., 1994). Methyl eugenol
produced sister chromatid exchange (SCE) in Chinese hamster ovary (CHO) cells,
but only in the presence of metabolic activation and at near-cytotoxic levels.
Therefore, the reported positive findings were likely due to cytotoxicity that resulted
in release of lysosomal nucleases, which may have led to a false-positive response.
Safrole concentrations of 0.01–810 μg/ml did not produce increases in SCE
in CHO cells, human peripheral lymphocytes or Chinese hamster V79 cells (Evans
& Mitchell, 1981; Natarajan & van Kesteren-van Leeuwen, 1981; Perry & Thomson,
1981; Douglas et al., 1985; Lane et al., 1985; Obe et al., 1985; van Went, 1985).