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of the Msp I-digested DNA fragments was comparable in both control and chemically-exposed animals. Thus because m5dC was morę liable to deamination than dC, con-seąuently to m5dC —> T transition mutation (Holliday, 1993), it seems unłikely that 2-butoxyethanol induces any mutation by dC methylation.

3.2. Methylation Level ofthe Whole Genomes ofRats and Mice and Hydrophilic Adduct Determination in Mice

In addition to the Ha -ras genes’ methylation assessments, the overall methylation level of the DNA under subchronic or acute treatment performed on transgenic mice or rats was also investigated. The results are shown in Figurę 2A and 2B for the spleen DNA. The methylation pattem did not vary with either subchronic or acute 2-butoxyethanol exposure, thus suggesting that the Chemical did not induce any changes in the level of DNA methylation. The same pattem was found in all other organs evaluated. The mean values of m5dC content of DNA from mice and rats revealed no significant variation in rats (brain, kidney, liver, spleen and testis) and mice (brain, bonę marrow, spleen and testis) (Table I). These results led to the conclusion that the methylation of the whole genome was neither affected by acute nor by subchronic 2-butoxyethanol exposures.

In order to check for other modified nucleotides which could carry aliphatic side chains, the autoradiograms of the chromatograms in which the m5dC contents were detected were overexposed. The modified nucleotides mentioned (i.e. hydrophilic adducts which should migrate in the neighborhood of the major nucleotides, but at lev-els far below those of the latter) could be due to 2-butoxyethanol metabolites after detoxification. Indeed it is known that 2-butoxyacetaldehyde is one such metabolite

Table I Methylation levels of the DNA extracted from different organs originating from animals treated with 2-butoxyethanol compared to control animals. The results are expressed in percentages of m5dC related to total cytidylic acid derivatives (m5dC + C). They correspond to the mean values of 5 assays (mice) and 15 assays (rats) performed on each type Of DNA. No differences between groups were found.

MICE (MALES)

	CONTROL					TREATED
	Day 0	120	10		15	60		120
Brain	3.0	3.1	2.9		3.1	3.3	3.2	3.5
Bonę marrow	3.9	3.3	4.0		3.9	3.8	4.3	3.5
Spleen	5.2	5.1	5.2		4.8	5.2	5.5	5.5
Testis	3.9	3.8	3.7		3.9	4.1	3.9	3.9
		MICE (FEMALES)					RATS (MALES)
	CONTROL			TREATED		CONTROL		TREATED
	Day 60	120	10	60	90	120	Day 1	Day 1
Brain	4.2	3.7	3.1	3.9	3.6	3.6	3.5	4.0
Bonę marrow	4.1	4.1	5.2	3.7	4.0	3.7	ND	ND
Spleen	4.6	5.1	4.6	4.6	5.8	5.5	3.6	3.9
Testis	-	-	-	-		-	3.4	3.7
Liver	ND	ND	ND	ND	ND	ND	3.5	3.8
Kidney	ND	ND	ND	ND	ND	ND	4.0	3.6

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