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throcytes (MNPCE) peaked 24 h after treatment at the low and middle dose levels, but at the high dose level the peak incidence of MNPCE was observed 48 h after treatment. In the double treatment with a 24-h inte~'al, the incidences of MNRET and MNPCE peaked 24 h after the second treatment in each strain. To study the enhancement effect of the multiple treatments in MN induction, 2-AAF was administered once or twice one week before starting the MN test. No effect of the pretreatment was observed.

8 Association of Microbial Mutation Testing Labo- ratory, C.O. Japan Bioassay Laboratory, 2445 Hi- rasawa, Hadano 257 (Japan)

Collaborative studies on the sensitivity of Salmo- nella tester strains

The sensitivities of Salmonella TA98 and TAI00 were studied on reference and laboratory strains at the same time by the plate incorpora- tion and preincubation methods at 20 laborato- ries. Reference strains of the same lot of precul- ture were divided into 200-/zl aliquots and frozen in vials. Several vials were distributed to each participating laboratory. Preculture was done by inoculating both reference and laboratory strains at fixed volumes into nutrient broth after thawing frozen cultures culture conditions were each lab- oratory standard techniques. Dose-response curves were obtained by two tests on four fixed doses of AF.2, which was also distributed to all laboratories from the same lot of material, with- out metabolic activation. Solvent control (HaP and DMSO) and background control with only biotin were measured.

Most laboratories showed no difference in spe- cific mutagenic activity of AF-2 between refer- ence and laboratory strains. However, the varia- tion between laboratores was 5-6 times. The variation between laboratories seemed due to testing and preculture conditions, not to strains. The solvent control values of TAI00 tended to increase with preculture time and the sensitivity of TA98 to AF-2 was strongly influenced by the preculture condition.

9 Atai, H., Y. Hatakeyama and S. Suzuki, Preclini- col Research Laboratories, Central Institute for Experimental Animals, Miyamae-ku, Kawasaki 216 (Japan)

Effects of benz[a]anthracene on the micronu- cleus test in mice

Benz[a]anthracene (BA) has been categorized as a member of group 2A (probable human car- cinogen) by the International Agency for Re- search on Cancer. However, data on micronu- cleus tests using BA are seldom found. Thus, in order to investigate the effect of BA, a micronu- cleus test was conducted in male CD-1 and BDF1 mice after intraperitoneal administration of BA. An increase in micronucleated polychromatic ery- throcytes in the bone marrow or micronucleated peripheral reticulocytes (MNRETs) was not ob- served after the administration of BA at up to 1000 mg/kg in either strain of mouse.

Conjugation with glutathione (GSH) serves to inactivate the reactive metabolite of polycyclic aromatic hydrocarbons (PAH) such as BA. Thus, the micronucleus test with BA was then carried out using OSH-depleted mice which were pre- treated with buthionine sulfoximine or phorone. The frequency of MNRETs increased after in- traperitoneal administration of BA at I000 mg/kg. These findings suggest that in GSH-de- pleted mice, it is possible to detect even slight carcinogenic activity of weak PAil, by observing the incidence of micronuclei formed.

This study was conducted as part of the sixth collaborative study by the Collaborative Study Group for the Micronucleus Test.

I0 Awogi, T., and Y. Ohara, Otsuka Pharmaceutical Co. Ltd., 463-10, Kagasuno, Kawauehi-cho, Tokushima 771-01 (Japan)

A limitation of rat peripheral blood reticulocytes for the micronucleus assay

Hayashi et al. reported that peripheral blood reticulocytes (PB-RETs) of rats were useful to