Abstract
Sphingoid bases, such as unsaturated sphingosine (So) and its corresponding dihydro-saturated species sphinganine (Sa), are present in cell samples in low abundance. This fact combined with their low-to-moderate electrospray ionization (ESI) potential, compared to other sphingolipids such as sphingomyelins, limits their detection and quantitation by liquid chromatography-tandem mass spectrometry (LC-MS(2)). To enhance the ESI efficiency of sphingoid bases, a novel procedure to generate stably derivatized analytes that enhance the LC-MS(2) detection of sphingoid bases when analyzed using LC-MS(2) was developed. In this method, a ruthenium complex, [4-(N-succimidyloxycarbonyl propyl)-4'-methyl-2,2'-bipyridine] bis(2,2'-bipyridine) Ru(II) dihexafluorophosphate, is added directly to a cell extract. This complex reacts with and covalently binds to an amino group within the sphingoid bases. The dicationic nature of the ruthenium ion enhances the compound's ionization efficiency resulting in increased LC-MS(2) signals for the derivatized sphingoid bases. Consequently, the detection and quantitation of sphingoid bases are greatly improved.