Development of environmental low-load-type analytical system of marine lipids

本研究的目的是开发一种环境低负荷型的高效液相色谱分析系统，以尽可能减少有机溶剂的消耗。为此，使用常规的ODS柱(250×4.6 mm×i.d.)进行了高效液相色谱-电喷雾电离-质谱仪(ESI-MS)和高效液相色谱-荧光检测。手性相高效液相色谱和反相高效液相色谱-紫外检测也使用了浆液填充的熔融二氧化硅毛细管(40 cm×0.1-0.3 mm i.d.)。分别与各种手性固定相和ODS柱联用。采用正己烷-1，2-二氯乙烷-乙醇三元流动相对手性相高效液相色谱法进行了优化。利用这些技术，我们成功地分离了甘油脂的分子种类(一、二、三酰甘油、烷基甘油、磷脂酰胆碱、磷脂酰乙醇胺、磷脂酰甘油、磷脂酰肌醇)，包括对映异构体和非对映异构体。甘油脂被层析为完整的形式、3，5-二硝基苯基氨基甲酸酯和1-苯甲基氨基甲酸酯。在本研究中，我们还实现了甘油脂的反相异构体的分离和鉴定，如20：5-16：0和22：6-16：0的1，2-二酰基甘油。本研究提供了一种环保的低负载型海洋油脂分析方法，与传统的高效液相色谱方法相比，可以减少有机溶剂的消耗。

The aim of this study was to develop an environmental low-load-type analytical system of marine lipids using high-performance liquid chromatography (HPLC), which reduces a consumption of organic solvents as much as possible. For this purpose, HPLC in conjunction with electrospray ionization mass spectrometry (ESI-MS) and HPLC with fluorescent detection were done using conventional ODS columns (250 x 4.6mm x i.d.). Chiral-phase HPLC and reversed-phase HPLC with UV detections were also applied using slurry-packed fused-silica capillaries (40cm x 0.1-0.3mm i.d.) with a various kind of chiral stationary phases and ODS, respectively. The chiral-phase HPLC analysis was optimized by using a ternary mobile phase, n-hexane-1,2-dichloroethane-ethanol. Using these techniques, we obtained successful separations of the molecular species of the glycerolipids (mono-, di-, triacylglycerols, alkylglycerols, phosphatidylcholines, phosphatidyl-ethanolamines, phosphatidylglycerols, phosphatidylinositols) including enantiomers and diastereomers. The glycerolipids were chromatographed as intact forms, 3,5-dinitrophenylurethanes and 1-anthrylurethanes. In the present study, we also achieved the separation and identification of the reverse isomers of glycerolipids, such as 1,2-diacylglycerols of 20:5-16:0 and 22:6-16:0, using reversed-phase HPLC/ESI-MS. The present study provides environmental low-load-type analytical methods of marine lipids, which can reduce a consumption of organic solvents compared to conventional HPLC methods.