A Tandem Mass Spectrometric Approach for Determining the Structure of Molecular Species of Ceramide in the Marine Sponge,Haliclona Cribricutis

Ceramides are important intracellular second messengers that play a role in the regulation of cell growth, differentiation and programmed cell death. Analysis of these second messengers requires a sensitive and specific analytical method to detect individual ceramide species and to differentiate between them. Eight molecular species of ceramide were identified from the marine sponge Haliclona cribricutis using electrospray ionization tandem mass spectrometry (ESI-MS/MS). From this marine sponge, N-hencicosanoyl (N21:0) to N-hexasanoyl (N26:0) octadecasphing-4 (E)-enine have been reported for the first time. The ESI-MS spectra gave several strong protonated [M+H]⁺ molecular ions with the corresponding bis (2-ethyl hexyl) phthalate adduct [M+H+DHEP]⁺. The collision-induced dissociation on ceramides at m/z 622.7337, 636.7645, 650.7789, 664.7925 and 678.8130 conducted at low-collision energy produced product ions with good characteristic at m/z 252.31, 264.32, 278.33, 282.33 and 296.35 for d18: 1 sphingosine, regardless of the length of the fatty chain. The MS/MS of the phthalate adduct [M+H+DHEP]⁺ at m/z 1013.1820, 1027.1971, 1041.2176, 1055.2394 and 1069.2573 also yielded characterizing product ions for sphingosine and confirmed the molecular ion at m/z 391 for bis (2-ethyl hexyl) phthalate. The major ions in the [M+H]⁺ and [M+H+DHEP]⁺ were due to neutral loss of [M+H–H₂O]⁺ and [M+H–(H₂O)₂]⁺.