Normal- and reverse-phase HPLC separations of fluorescent (NBD) lipids

✍ Scribed by Ona C. Martin; Richard E. Pagano

• Publisher: Elsevier Science
• Year: 1986
• Tongue: English
• Weight: 744 KB
• Volume: 159
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(86)90313-1

✦ Synopsis

We have developed two high-performance liquid chromatography methods for separating a number of fluorescent 4-nitrobenzo-2-oxa-1,3diazole

(NBD) analogs of glycerolipids and sphingolipids. Samples of fluorescent lipid analogs containing NBDaminocaproyl (C6-NBD) or NBDaminododecanoyl (&NBD) acyl chains were synthesized and analyzed by the following HPLC methods. (i) An isocratic normal-phase method permitted resolution of a mixture of the 1,2-(palmitoyl, C6-NBD)-analogs of triacylglycerol, diacylglycerol, phosphatidic acid, phosphatidylethanolamine, and phosphatidylcholine in less than 10 min, while a mixture of the (Cs-NBD)labeled analogs of ceramide, glucocerebroside, and sphingomyelin was separated in approximately 15 min. This method also detected various (C6-NBD)-phosphatidylcholine and -phosphatidylethanolamine molecules which differed only in their nonfluorescent acyl (oleoyl or palmitoyi) chains, and readily separated nonfluorescent dipalmitoylphosphatidylcholine from both (C6-NBD)-and (C,,NBD)-phosphatidylcholine derivatives. (ii) An isocratic reverse-phase system permitted separation of isomers of fluorescent phosphatidylcholine, -ethanolamine, -glycerol, -inositol, -serine, and phosphatidic acid in which the NBD-fatty acid was present in either the sn-1 or sn-2 position of the glycerol backbone. 0 1986 Academic press, IIIC.