Kunihiko Ogawa

In vitro biotransformation of atrazine by rat liver microsomal cytochrome P450 enzymes

We studied atrazine (ATZ) metabolism in male and female rat liver microsomes in vitro, and the major metabolite was deisopropylatrazine (DeiPr-ATZ) with deethylatrazine (DeEt-ATZ) and 1-hydroxyisopropylatrazine (iPrOH-ATZ) as minor metabolites in both sexes. The enzyme kinetics of ATZ biotransformation were examined by means of Eadie-Hofstee analyses. Although no remarkable sex difference of Michaelis Menten values for each pathway was observed, Cl(int)S (Vmax/Km) for DeiPr-ATZ, DeEt-ATZ and iPrOH-ATZ were slightly higher in female than in male rats. The formation of DeiPr-ATZ, DeEt-ATZ and iPrOH-ATZ from ATZ was substantially inhibited by SKF-525A, metyrapone, diallyl sulfide, 7-ethoxycoumarin, benzphetamine, nicotine, testosterone and lauric acid in both sexes. Cimetidine effectively inhibited the formation of all metabolites in male rats. On the other hand, the inhibition rates of the formation of DeiPr-ATZ and iPrOH-ATZ by cimetidine in female rats were lower than those in male rats, and DeEt-ATZ was hardly affected by the chemicals. In contrast with the results for cimetidine, the inhibition of ATZ biotransformation by bufuralol was more effective in female than in male rats. Anti-rat CYP2B1 and CYP2E1 antibodies effectively inhibited DeiPr-ATZ, DeEt-ATZ and iPrOH-ATZ formations in both sexes. Anti-rat CYP2C11 antibody also inhibited the three metabolites in both sexes, with the inhibition rates higher in male than in female rats, similar to cimetidine. In the case of anti-rat CYP2D1 antibody, the inhibitory effect on ATZ biotransformation in male rats was less than that in female rats. On the other hand, anti-rat CYP1A2, CYP3A2 and CYP4A1 antibodies did not affect the ATZ biotransformation in either sex. There was no significant correlation between the formation rate of ATZ metabolites and P450 isoform levels in either sex. These results may mean that CYP2B2, CYP2C11, CYP2D1 (only iPrOH-ATZ formation) and CYP2E1 in male rats, and CYP2B2, CYP2D1 and CYP2E1 in female rats are involved ATZ metabolism in liver, and that the substrate specificity of P450 isoforms for ATZ is broad.

Determination of water in substances containing active chlorine by Karl-Fischer titration

カールフィシャー法による水分測定において,遊離塩素あるいは活性塩素はカールフィシャー試薬と水との反応生成物であるヨウ化水素を酸化してヨウ素を遊離する妨害を示す.この妨害を抑制するのに,遊離塩素あるいは活性塩素を含む物質をあらかじめピリジン-二酸化イオウのメタノール溶液で処理することにより,塩素が不活性化することを確かめた.遊離塩素約1%を含む四塩化炭素,1,2-ジクロルエタソおよび活性塩素を含むジクロルイソシアヌール酸カリウムを,ピリジン-二酸化イオウのメタノール溶液で処理してから滴定することにより,試料中の0.001～0.1%の微量水分を正確に定量できた.またアリルスルホン酸塩化物も同じ方法で処理すれば容易に水分が測定できることを確かめた.