クロラムフェニコ-ル耐性菌によるクロラムフェニコ-ルのアセチル化機構--GC-MASSによる解析
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Two types of diacetylated chloramphenicol 1, 3-O-O-di [1-12C] acetyl chloramphenicol and 1, 3-O-O-di [1-14C] acetyl chloramphenicol, were subjected to GC-mass analysis. The former showed the presence of prominent mass peaks at m/e 195, 212, 214 and 216, and the latter at m/e 197, 214, 216 and 218. Thus, the fragment ions of the compound diacetylated with. [1-14C] acetyl CoA were detected at a higher mass field by m/e 2 than those of the one diacetylated with [1-12C] acetyl CoA. The C1-C2 bond of diacetylated chloramphenicol was subjected to cleavage. The mass peaks at m/e 195 and 197 and those at m/e 212, 214, 216 and 218 were produced by the fragment containing C-1 and that containing C-3, respectively. The technique was applied to identify the acetoxy group attached to C-1 or C-3 of diacetylated chloramphenicol.To clarify the exact position acetylated by chloramphenicol acetyltransferase in chloramphenicol, the [1-14C] acetyl group wes introduced into 3-O-[1-12C] acetyl chloramphenicol, by using [1-14C] acetyl CoA. The mass peaks of diacetylated chloramphenicol appeared at m/e 195, 212, 214, 216 and 218. Especially, the relative intensity at m/e 214 increased. No peaks occurred, however, at m/e 197. The result described above indicated that the [1-12C] acetyl group was attached to C1-OH and the [1-14O] or the [1-12C] acetyl group to C3-OH in chloramphenicol.From these result and those of kinetic studies, it was considered that both mono- and diacetylation or chloramphenicol by chloramphenicol acetyltransferase might take place in the hydroxy group attached to C-3 of chloramphenicol.The presence of the [1-14C] acetyl group in monoacetylated chloramphenicol suggested a mutual exchange between the [1-12C] acetyl and the [1-14C] acetyl groups.