Swaffar D S, Harker W G, Pomerantz S C, Lim H K, Yost G S
Department of Pharmacology and Toxicology, University of Utah, Salt Lake City 84112.
Drug Metab Dispos. 1992 Sep-Oct;20(5):632-42.
We have previously reported that the antineoplastic agent, procarbazine, in aqueous solutions was chemically oxidized to its azoxy metabolites (methylazoxy and benzylazoxy). To determine if there was additional metabolism of the most active metabolite, methylazoxyprocarbazine, it was incubated in the presence and absence of CCRF-CEM human leukemia cells. Incubations were extracted, and potential metabolites were detected by HPLC with UV detection and by combined HPLC and thermospray mass spectrometric analysis. The major metabolite identified by HPLC with UV detection of the extracts was N-isopropyl-p-formylbenzamide; this was identified by comparison of its retention time with that of a synthesized standard. This identification was further corroborated by HPLC/thermospray mass spectrometry (LC/MS). Analysis of the extracts by LC/MS also showed the presence of a closely eluting peak that had a protonated molecular ion at m/z 207. This new metabolite was identified as N-isopropyl-(benzene-1,4-bis-carboxamide) by 1H NMR and gas chromatography/ion trap mass spectrometry. This metabolite is postulated to arise from breakage of the N-N bond in the hydrazine portion of the molecule. Reconstructed ion (m/z 236) current profiles from the analysis of the cell extracts indicated that there was only a trace amount of methylazoxyprocarbazine left after a 72-hr incubation. Interestingly, a peak with the same molecular weight as the parent compound (methylazoxyprocarbazine) was observed in the cellular incubations and also in extracts of control incubations in which methylazoxyprocarbazine was incubated in medium without cells. This unknown was silylated and identified as a hydroxyazo compound by an ion trap mass spectrometer operated under both single and multiple-stage mass analysis. Formation of this decomposition product appears to involve a novel intramolecular rearrangement of methylazoxyprocarbazine in solution. This pathway may be responsible for the formation of the ultimate cytotoxic species by chemical decomposition of procarbazine.
我们之前曾报道，抗肿瘤药物丙卡巴肼在水溶液中会被化学氧化为其偶氮氧化代谢产物（甲基偶氮氧化和苄基偶氮氧化产物）。为了确定最具活性的代谢产物甲基偶氮氧化丙卡巴肼是否还有其他代谢情况，我们在有和没有CCRF - CEM人白血病细胞存在的条件下对其进行孵育。孵育后进行萃取，通过带有紫外检测的高效液相色谱法（HPLC）以及HPLC与热喷雾质谱联用分析来检测潜在的代谢产物。通过对萃取物进行紫外检测的HPLC鉴定出的主要代谢产物是N - 异丙基 - 对甲酰基苯甲酰胺；通过将其保留时间与合成标准品的保留时间进行比较来鉴定。高效液相色谱/热喷雾质谱法（HPLC/TS/MS）进一步证实了这一鉴定结果。通过LC/MS对萃取物进行分析还显示存在一个与之紧密洗脱的峰，其质子化分子离子的质荷比为m/z 207。通过¹H核磁共振以及气相色谱/离子阱质谱法将这种新的代谢产物鉴定为N - 异丙基 -（苯 - 1,4 - 二羧酰胺）。据推测，这种代谢产物是由于分子中肼部分的N - N键断裂而产生的。对细胞萃取物分析得到的重构离子（m/z 236）电流图谱表明，经过72小时孵育后仅残留痕量的甲基偶氮氧化丙卡巴肼。有趣的是，在细胞孵育物以及对照孵育物（即在无细胞的培养基中孵育甲基偶氮氧化丙卡巴肼）的萃取物中都观察到了一个与母体化合物（甲基偶氮氧化丙卡巴肼）分子量相同的峰。将这个未知物进行硅烷化处理，并通过在单级和多级质量分析下运行的离子阱质谱仪鉴定为一种羟基偶氮化合物。这种分解产物的形成似乎涉及甲基偶氮氧化丙卡巴肼在溶液中的一种新型分子内重排。这条途径可能是丙卡巴肼通过化学分解形成最终细胞毒性物质的原因。
