Yu Keshun, Liu Huazhen, Kachroo Pradeep
Department of Plant Pathology, University of Kentucky, Lexington, Kentucky, USA.
Bio Protoc. 2020 Dec 5;10(23):e3841. doi: 10.21769/BioProtoc.3841.
Pipecolic acid (Pip), a non-proteinacious product of lysine catabolism, is an important regulator of immunity in plants and humans alike. For instance, Pip accumulation is associated with the genetic disorder Zellweger syndrome, chronic liver diseases, and pyridoxine-dependent epilepsy in humans. In plants, Pip accumulates upon pathogen infection and is required for plant defense. The aminotransferase ALD1 catalyzes biosynthesis of Pip precursor piperideine-2-carboxylic acid, which is converted to Pip via ornithine cyclodeaminase. A variety of methods are used to quantify Pip, and some of these involve use of expensive amino acid analysis kits. Here, we describe a simplified procedure for quantitative analysis of Pip from plant tissues. Pipecolic acid was extracted from leaf tissues along with an internal standard norvaline, derivatized with propyl chloroformate and analyzed by gas chromatography-coupled mass spectrometry using selective ion mode. This procedure is simple, economical, and efficient and does not involve isotopic internal standards or multiple-step derivatizations.
哌啶酸（Pip）是赖氨酸分解代谢的一种非蛋白质产物，在植物和人类中都是重要的免疫调节因子。例如，Pip的积累与人类的遗传性疾病泽尔韦格综合征、慢性肝病和吡哆醇依赖性癫痫有关。在植物中，病原体感染时Pip会积累，并且是植物防御所必需的。转氨酶ALD1催化Pip前体哌啶-2-羧酸的生物合成，后者通过鸟氨酸环脱氨酶转化为Pip。有多种方法用于定量Pip，其中一些方法涉及使用昂贵的氨基酸分析试剂盒。在此，我们描述了一种从植物组织中定量分析Pip的简化程序。哌啶酸与内标正缬氨酸一起从叶片组织中提取，用氯甲酸丙酯衍生化，并使用选择性离子模式通过气相色谱-质谱联用进行分析。该程序简单、经济且高效，不涉及同位素内标或多步衍生化。
