Fundamental and Molecular Mechanisms of Mutagenesis

ELSEVIER Mutation Research 306 (1994) 107-109

Series: C u r r e n t Issues in M u t a g e n e s i s and Carc inogenes i s , No. 44

Benzyl acetate: from mutagenic carcinogen to non-mutagenic non-carcinogenic in 7 years?

J o h n A s h b y *

Zeneca Central Toxicology Laboratory, Alderley Park, Macclespeld, Cheshire SKIO 477, UK

(Received 4 January 1994; accepted 6 January 1994)

Key words: Current issues; Benzyl acetate

In 1986 the US NTP reported benzyl acetate to be carcinogenic to mice and rats (Anon., 1986). Genetic toxicity experiments reported at the same time defined benzyl acetate as a 'unique gene mutagen' by virtue of its isolated activity in the mouse lymphoma assay (Table 1). Such data con- tributed to the idea that chemicals should be assessed separately for their ability to induce chromosomal or gene mutations in mammalian cells in vitro (discussed at the time in Ashby, 1988).

In the 1986 NTP Technical Report (Anon., 1986) the possibility was raised that the corn oil vehicle might have contributed to the observed male rat pancreatic tumours. Further, the fact that benzyl acetate is a stomach irritant when dosed by gavage to the mouse raised the possibil- ity that this activity had influenced the induction of the mouse stomach tumours. Mindful of these possible complicating factors, and of the fact that benzyl acetate is used as a flavouring agent in food, the NTP re-evaluated the carcinogenicity of

* Corresponding author.

this agent in a feed study (Anon., 1993). It was found to be non-carcinogenic to both species of rodent (Table 1).

During the course of the repeat cancer bioas- say the genetic toxicity database on benzyl ac- etate was considerably extended (Anon., 1993; see Table 1). The positive mouse-lymphoma re- sponse observed in the first study was confirmed and extended to tests conducted in the absence of $9 mix. All other in vitro and in vivo genetic toxicity test results were negative. These negative results were extensive and included assessment of the peripheral blood for micronucleated erythro- cytes in mice exposed to the heroic dose of 50 000 ppm in diet, equivalent to approximately 9 g/kg bodyweight per day of benzyl acetate, for 13 weeks.

In 1986, benzyl acetate was considered a ro- dent carcinogen that was only detected as geno- toxic in the mouse-lymphoma assay. Now, it is reported to be a rodent non-carcinogen that is supported by extensive negative genotoxicity data, yet it remains mutagenic in the mouse-lymphoma assay. The changing fortune of benzyl acetate is pertinent to recent discussion in Current Issues of how best to test chemicals for genetic toxicity and

0027-5107/94/$07.00 © 1994 Elsevier Science B.V. All rights reserved SSDI 0 0 2 7 - 5 1 0 7 ( 9 4 ) 0 0 0 0 9 - T

108 J. Ashby/Mutation Research 306 (1994) 107-109

carcinogenic potent ia l (Ashby, 1993; Madle, 1993; Zeiger, 1994).

A n in teres t ing aspect of these two sets of carcinogenici ty bioassay results is that the peak blood levels of benzoic acid (the pr imary metabo-

lite of benzyl acetate) were up to 300 t imes higher following gavage dosing than in the feed studies (Anon. , 1993). While this may go some way to-

wards explaining the different bioassay outcomes it is pe r t inen t to note that dietary in take is the major route of h u m a n exposure, the o ther route being by skin exposure to lotions and perfumes. Three quest ions are raised by these carcinogenic- ity data. First, how should the route of exposure in roden t -cancer bioassays be selected and ratio- nal ized? Second, how could low-dose risk estima- t ions be adjusted for differences in peak blood levels caused by changes to the route of expo- sure? Third, what is the presen t carc inogenic

status of benzyl acetate? The last of these three quest ions will doubtless be discussed by those in teres ted in the predictive value of the mouse-

lymphoma assay, and by those concerned with the potent ia l carc inogenic hazard presen ted by expo- sure of humans to benzyl acetate. It would seem that the positive mouse- lymphoma data are severely isolated, and that this presents a ' local ' p rob lem for genet ic toxicologists to solve. A ny residual discussion of the possible carc inogenic hazard posed to h u m a n s by exposure to benzyl acetate must be related solely to the fact, as highlighted in the N TP report , that about half a mil l ion people in the U n i t e d States were exposed to this material , in one way or another , be tween 1981 and 1983 (Anon. , 1993). Nonetheless , the data shown in Table 1 are most consis tent with benzyl acetate not posing a carcinogenic hazard to humans .

Table 1 Summary of genetic toxicity and carcinogenicity data presented in two NTP Technical Reports on benzyl acetate

Assay NTP TR 250 (1986) NTP TR 431 (1993)

Salmonella - - CHO SCE - - CHO CA - -

Mouse-lymphoma + ( + $9 only) + ( + $9) L5178Y tk ÷/ (TFT)

Drosophila SLRL Mouse BM SCE Mouse BM CA Mouse BM MN Mouse per. blood MN

Carcinogenicity tests F344 rat, gavage in

corn oil (500 mg/kg) B6C3F1 mouse,

gavage in corn oil (1000 mg/kg)

F344 rat, feed study (12000 ppm;

550 mg/kg) B6C3F1 mouse, feed

study (3000 ppm; ~ 350 mg/kg)

+

+

Comment

Extra trial (_+ $9) added to 1993 report Same data in each report Extra two trials ( - $9) and one more

+ $9 in 1993 report 1986 data supplemented with additional

+ $9 repeat. Also, 3 studies ( - $9) positive in a different laboratory

Feed and injection Tested to 1700 mg/kg Tested to 1700 mg/kg Tested to 1250 mg/kg 13 wk at 50000 ppm in diet (~ 8000 mg/kg males;

~ 9000 mg/kg females)

Male rate pancreatic acinar cell adenomas

Male and female mouse liver adenomas and forestomach adenomas/carcinomas

Dose levels set by palatability. Higher levels could have been employed in the rat study

Many sites of non-neoplastic lesions. MTD achieved

CA, chromosomal aberrations; BM, bone marrow; MN, micronucleus. The mouse peripheral blood MN assays were done on animals from the 13-week bioassay dose-setting study. MTD, maximum tolerated dose.

J. Ashby/Mutation Research 306 (1994) 107-109 109

1. References

Anon. (1986) Toxicology and carcinogenesis studies of benzyl acetate, NTP Technical Report 250.

Anon. (1993) Toxicology and Carcinogenesis studies of benzyl acetate, NTP Technical Report 431.

Ashby, J. (1988) Discussion Forum, Mutagenesis, 3, 463-465.

Ashby, J. (1993) Precedents or Possibilities, Mutation Res., 298, 291-295.

Madle, S. (1993) Problem of the harmonization of philoso- phies of genotoxicity testing, Mutation Res., 300, 73-76.

Zeiger, E. (1994) Strategies and philosophies of genotoxicity testing: What is the question? Mutation Res., 304, 309-314.