In vitro and in vivo evaluations of the methaemoglobinaemic potential of xylidine isomers in the rat.

The objective of the present study was to evaluate the methaemoglobinaemic potential of the six isomers of xylidine (XYL) by two approaches: in vitro, using rat red blood cells and hepatic post-mitochondrial fractions in a two-compartmental dialysis system; and in vivo, following a single oral dose of 4.8 mmol kg(-1) (p.o.) of each of the six XYL isomers. The in vitro experiments showed that all six XYL isomers at 1 mM concentration induced significant methaemoglobinaemia in the presence of active hepatic fractions, whereas non-bioactivated XYL isomers were totally inactive. At lower incubation concentrations (0.3 mM and 0.06 mM), 3,5-XYL was still active, whereas the other isomers were less potent (0.3 mM) or totally ineffective (0.06 mM). The in vivo experiment revealed that all XYL isomers, except 3,5-XYL, did not induce significant methaemoglobinaemia after a single oral dose of 4.8 mmol kg(-1). The maximal percentage of methaemoglobin was 31.3 +/- 1.5 in the 3,5-XYL-treated rats, whereas it never exceeded 3% in all the other treatment groups, indicating that 4.8 mmol kg(-1) (p.o.) is in fact a no-observable-adverse-effect level for these XYL isomers. The quantitative differences between in vivo and in vitro results may have been due to additional bioactivation pathways (N-hydroxylation or ring hydroxylation) mediated by high Km enzymes operative at the high incubation concentrations used in vitro. The results of the present study suggest that 3,5-XYL is likely to be the only active isomer in the Sprague-Dawley rat at low exposure levels.