Kondo Y, Kawai Y, Hayashi T, Ohnishi M, Miyazawa T, Itoh S, Mizutani J
Plant Ecochemicals Project, Research Development Corporation of Japan, Hokkaido.
Biochim Biophys Acta. 1993 Nov 3;1170(3):301-6. doi: 10.1016/0005-2760(93)90014-z.
Lipoxygenase (LOX) activities in the crude extracts of soybean cotyledons on phospholipid were measured by a chemiluminescence (CL) assay system. The activity of LOX on phospholipid increased at 6 h after treatment with fungal elicitor, whereas the activity in the control experiment without elicitor treatment declined with incubation. A rise of LOX activity at 6 h incubation is considered to be induced by the elicitor treatment. The LOX fraction obtained from soybean cotyledons at 6 h after elicitation by a DEAE column chromatography was reacted with 1-palmitoyl-2-linoleoyl-phosphatidylcholine substrate. The reaction product was analyzed by HPLC equipped in series with a UV (205 nm) and a CL detector. Most of the substrate was converted to a hydroperoxy compound. By the CL-HPLC analysis after transesterification of reaction product and by mass spectrum of the trimethylsilyl derivative after reduction of methyl ester confirmed that the linoleic moiety was subjected to oxygenation at 13-position. The steric analysis of menthoxyacetyl derivative prepared from hydroxy fatty acid methyl ester showed S configuration. These findings suggest that the oxygenation of membrane lipids in biological plant tissues might be initiated by the direct action of LOX.
通过化学发光(CL)测定系统测量大豆子叶粗提物中脂氧合酶(LOX)对磷脂的活性。用真菌激发子处理6小时后,LOX对磷脂的活性增加,而在未用激发子处理的对照实验中,其活性随孵育时间下降。孵育6小时时LOX活性的升高被认为是由激发子处理诱导的。用DEAE柱色谱法从激发6小时后的大豆子叶中获得的LOX组分与1-棕榈酰-2-亚油酰磷脂酰胆碱底物反应。反应产物通过串联配备紫外(205nm)和CL检测器的HPLC进行分析。大部分底物转化为氢过氧化物。通过反应产物酯交换后的CL-HPLC分析以及甲酯还原后三甲基硅烷基衍生物的质谱分析证实,亚油酸部分在13位发生了氧化。由羟基脂肪酸甲酯制备的薄荷氧基乙酰衍生物的立体分析显示为S构型。这些发现表明,生物植物组织中膜脂的氧化可能是由LOX的直接作用引发的。