Structural distinction of diacyl-, alkylacyl, and alk-1-enylacyl glycerophosphocholines as [M - 15]⁻ ions by multiple-stage linear ion-trap mass spectrometry with electrospray ionization

Abstract

We describe a linear ion-trap (LIT) multiple-stage (MS(n)) mass spectrometric approach towards differentiation of alkylacyl, alk-1-enylacyl- and diacyl-glycerophoscholines (PCs) as the [M - 15]⁻ ions desorbed by electrospray ionization (ESI) in the negative-ion mode. The MS⁴ mass spectra of the [M - 15 - R²'CH = CO]⁻ ions originated from the three PC subfamilies are readily distinguishable, resulting in unambiguous distinction of the lipid classes. This method is applied to two alkyl ether rich PC mixtures isolated from murine bone marrow neutrophils and kidney, respectively, to explore its utility in the characterization of complex PC mixture of biological origin, resulting in the realization of the detailed structures of the PC species, including various classes and many minor isobaric isomers.

Figure 1

The high resolution (R =100,000 at m/z 400) ESI-MS spectrum of the [M + Cl]⁻ ions of p18:0/18:1-PC. The theoretical plot (using the software downloaded from http://www.alchemistmatt.com/) from the elemental composition (C₄₄H₈₆O₇NPCl) is shown in “inset,” which gives an identical profile

Figure 2

The MS² spectrum of the [M + Cl]⁻ ions of p18:0/18:1-PC at m/z 806.6 (a), its MS³ spectrum of the ion of m/z 756.6 (806→756) (b), the MS³ spectrum of the [M – 15]⁻ ion of a18:1/18:1-PC at m/z 756.6 (806→756) (c), the MS⁴ spectra of the ions of m/z 492 (806→756→492) arising from p18:0/18:1-PC (d), and from a18:1/18:1-PC (e) isomers

Figure 3

The MS³ spectrum of the [M – 15]⁻ ions of p18:0/20:4-PC standard at m/z 778.6 (828→778) (a), its MS⁴ spectrum at m/z 492 (828→778→492) (b), the MS³ spectrum of the [M – 15]⁻ ions of a16:0/20:4-PC standard at m/z 752.5 (802→752) (c), its MS⁴ spectrum at m/z 466 (802→752→466) (d), the MS³ spectrum of the [M – 15]⁻ ions of 16:0/18:1-PC standard at m/z 744.6 (794→744) (e), and its MS⁴ spectra of the ions of m/z 480 (794→744→480) (f)

Figure 4

The MS³ spectrum of the [M – 15]⁻ ions of m/z 728.5 (a), its MS⁴ spectra of the ions of m/z 490 (778→728→490) (b), MS³ spectrum of the [M – 15]⁻ ions of 756.6 (c), its MS⁴ spectra of the ions of m/z 494 (806→756→494) (d), and 518 (806→756→518) (e) arising from the PC extract of bone marrow neutrophils

Figure 5

The MS³ spectrum of the [M – 15]− ions of m/z 776.5 (a), its MS⁴ spectra of the ions of m/z 466 (826→776→466) (b), and of m/z 490 (826→776→490) (c) arising from the PC extract of mouse kidney

Scheme 1

Fragmentation pathways (MSn) proposed for alkyl ether PC ([M – 15]⁻ ions)