Table 1. ESI-MS MS/MS MRM and HPLC conditions for each sphingolipid.
Seg | Compound | RT (min) | Pre-ion (m/z) | Pro-ion (m/z) | Fragmentor (V) | CE (eV) | ISTD |
LC-MS/MS run 1: For sphingosine, sphingosine-1-phosphate, ceramides, ceramides-1-phosphate | |||||||
1 | Sph (d17∶1) | 2.5 | 286.0 | 267.8 | 100 | 7 | ISTD |
Sph (d18∶1) | 2.7 | 300.3 | 282.9 | 90 | 3 | Sph (d17∶1) | |
Sph (d17∶1)-1-P | 4.3 | 365.9 | 250.0 | 100 | 11 | ISTD | |
Sph (d18∶1)-1-P | 5.1 | 380.3 | 264.1 | 110 | 12 | Sph (d17∶1)-1-P | |
Cer(d18∶1/2∶0) | 6.9 | 342.1 | 324.1 | 100 | 2 | ISTD | |
Cer(d18∶1/2∶0)-1-P | 8.7 | 422.0 | 264.1 | 100 | 29 | Cer(d18∶1/8∶0)-1-P | |
Cer(d18∶1/4∶0) | 9.0 | 370.2 | 352.1 | 100 | 2 | Cer(d18∶1/2∶0) | |
2 | Cer(d18∶1/6∶0) | 12.6 | 398.2 | 380.1 | 100 | 2 | Cer(d18∶1/10∶0) |
Cer(d18∶1/8∶0) | 15.7 | 426.3 | 408.2 | 140 | 3 | Cer(d18∶1/10∶0) | |
Cer(d18∶1/8∶0)-1-P | 17.2 | 506.0 | 264.1 | 120 | 26 | ISTD | |
Cer(d18∶1/10∶0) | 17.5 | 454.2 | 436.2 | 140 | 5 | ISTD | |
Cer(d18∶1/12∶0) | 19.4 | 482.4 | 464.2 | 140 | 7 | Cer(d18∶1/10∶0) | |
Cer(d18∶1/14∶0) | 21.5 | 510.4 | 492.1 | 120 | 6 | Cer(d18∶1/10∶0) | |
Cer(d18∶1/12∶0)-1-P | 21.5 | 562.1 | 264.1 | 120 | 26 | Cer(d18∶1/10∶0) | |
3 | Cer(d18∶1/16∶0) | 23.8 | 538.4 | 520.2 | 140 | 6 | Cer(d17∶1/18∶0) |
Cer(d18∶1/18∶1) | 24.6 | 564.4 | 546.2 | 130 | 6 | Cer(d17∶1/18∶0) | |
Cer(d17∶1/18∶0) | 25.1 | 552.4 | 250.1 | 130 | 30 | ISTD | |
Cer(d18∶1/18∶0) | 26.2 | 566.4 | 548.3 | 150 | 12 | Cer(d17∶1/18∶0) | |
Cer(d18∶1/16∶0)-1-P | 26.4 | 618.1 | 263.9 | 110 | 31 | Cer(d18∶1/8∶0)-1-P | |
Cer(d18∶1/18∶1)-1-P | 27.3 | 644.2 | 264.1 | 150 | 34 | Cer(d18∶1/8∶0)-1-P | |
4 | Cer(d18∶1/20∶0) | 28.6 | 594.4 | 576.2 | 140 | 8 | Cer(d17∶1/24∶1) |
Cer(d17∶1/24∶1) | 30.3 | 634.5 | 616.6 | 140 | 10 | ISTD | |
Cer(d18∶1/22∶0) | 31.3 | 622.5 | 604.2 | 140 | 8 | Cer(d17∶1/24∶1) | |
Cer(d18∶1/24∶1) | 31.4 | 648.7 | 630.2 | 140 | 12 | Cer(d17∶1/24∶1) | |
Cer(d18∶1/24∶0) | 33.8 | 650.7 | 632.3 | 140 | 8 | Cer(d17∶1/24∶1) | |
Cer(d18∶1/26∶1)* | 34.1 | 676.8 | 658.3 | 140 | 12 | Cer(d17∶1/24∶1) | |
Cer(d18∶1/26∶0)* | 36.6 | 678.8 | 660.3 | 140 | 8 | Cer(d17∶1/24∶1) | |
LC-MS/MS run 2: For dihydrosphingosine, dihydorsphingosine-1-phosphate, dihydroceramides and hexcosyl-sphingolipids | |||||||
1 | HexSph(d18∶1) | 3.2 | 462.1 | 282.2 | 130 | 21 | dhSph(d17∶0) |
dhSph(d17∶0) | 3.5 | 287.9 | 110.0 | 270 | 11 | ISTD | |
dhSph(d18∶0) | 4.2 | 302.1 | 284.2 | 120 | 5 | dhSph(d17∶0) | |
dhSph-1-P(d17∶0) | 6.4 | 367.8 | 110.0 | 270 | 9 | ISTD | |
dhSph-1-P(d18∶0) | 8.2 | 382.1 | 284.2 | 80 | 10 | dhSph-1-P(d17∶0) | |
dhCer(d18∶0/2∶0) | 12.0 | 344.1 | 326.2 | 110 | 10 | dhCer(d18∶0/8∶0) | |
HexCer(d18∶1/8∶0) | 15.9 | 588.2 | 264.2 | 100 | 36 | dhCer(d18∶0/8∶0) | |
dhCer(d18∶0/6∶0) | 16.0 | 400.1 | 382.2 | 130 | 10 | dhCer(d18∶0/8∶0) | |
dhCer(d18∶0/8∶0) | 17.5 | 428.2 | 410.1 | 150 | 15 | ISTD | |
2 | HexCer(d18∶1/12∶0) | 18.7 | 644.2 | 264.1 | 150 | 36 | Cer(d17∶1/18∶0) |
HexCer(d18∶1/16∶0) | 21.7 | 700.2 | 264.1 | 140 | 36 | Cer(d17∶1/18∶0) | |
HexCer(d18∶1/18∶1) | 22.2 | 726.1 | 264.2 | 160 | 36 | Cer(d17∶1/18∶0) | |
HexCer(d18∶1/18∶0) | 23.2 | 728.2 | 264.1 | 150 | 39 | Cer(d17∶1/18∶0) | |
dhCer(d18∶0/16∶0) | 23.4 | 540.2 | 522.4 | 160 | 18 | Cer(d17∶1/18∶0) | |
Cer(d17∶1/18∶0) | 23.7 | 552.4 | 250.1 | 130 | 30 | ISTD | |
dhCer(d18∶0/18∶1) | 24.0 | 566.2 | 548.4 | 140 | 18 | Cer(d17∶1/18∶0) | |
dhCer(d18∶0/18∶0) | 25.0 | 568.2 | 550.3 | 140 | 18 | Cer(d17∶1/18∶0) | |
3 | HexCer(d18∶1/24∶1) | 26.6 | 810.2 | 264.1 | 100 | 36 | Cer(d17∶1/24∶1) |
Cer(d17∶1/24∶1) | 27.4 | 634.5 | 616.6 | 140 | 10 | ISTD | |
dhCer(d18∶0/24∶1) | 28.6 | 650.3 | 632.5 | 150 | 24 | Cer(d17∶1/24∶1) | |
dhCer(d18∶0/24∶0) | 30.4 | 652.3 | 634.4 | 180 | 24 | Cer(d17∶1/24∶1) | |
LC-MS/MS run 3: For sphingomyelines | |||||||
1 | Lyso-SM(d18∶1) | 2.0 | 464.8 | 170 | 183.9 | 24 | Lyso-SM(d17∶1) |
Lyso-SM(d17∶1) | 2.0 | 450.8 | 160 | 183.9 | 24 | ISTD | |
SM(d18∶1/2∶0) | 3.6 | 507.3 | 183.9 | 160 | 24 | SM(d18∶1/17∶0) | |
SM(d18∶1/6∶0) | 4.9 | 563.3 | 183.9 | 100 | 24 | SM(d18∶1/17∶0) | |
2 | SM(d18∶1/12∶0) | 7.5 | 647.4 | 183.9 | 180 | 33 | SM(d18∶1/17∶0) |
SM(d18∶1/16∶0) | 9.0 | 703.5 | 184 | 160 | 30 | SM(d18∶1/17∶0) | |
SM(d18∶1/18∶1) | 9.3 | 729.4 | 183.9 | 190 | 33 | SM(d18∶1/17∶0) | |
SM(d18∶1/17∶0) | 9.4 | 717.5 | 184 | 100 | 30 | ISTD | |
SM(d18∶1/18∶0) | 9.8 | 731.6 | 183.9 | 190 | 33 | SM(d18∶1/17∶0) | |
SM(d18∶1/24∶1) | 11.9 | 813.6 | 183.9 | 240 | 30 | SM(d18∶1/17∶0) | |
SM(d18∶1/24∶0) | 13.0 | 815.6 | 183.9 | 200 | 33 | SM(d18∶1/17∶0) |
Seg, segment; RT, retention time; Pre-ion, precursor ion; Pro-ion, product ion; CE, collision energy; ISTD, internal standard.
Cer(d18∶1/26∶1) and Cer(d18∶1/26∶0) were quantitated using the calibration curve of the closest counterpart when commercial standards were not available. Cer(d18∶1/26∶1) ∼Cer(d18∶1/24∶1) and Cer(d18∶1/26∶0) ∼ Cer(d18∶1/24∶0).