HPLC Separation of D-sphingosine and related compounds on Ascentis^® Express C18

column

Ascentis Express C18, 10 cm X 2.1 mm, 2.7um particles (53823-U)

mobile phase

(A) Water; (B) Acetonitrile, each with 0.1% formic acid/10 mM ammonium formate

gradient

70% B for 8 min, 0.5 min to 90%B, hold for 1.5 min, back to 70% in 0.5 min, hold for 3.5 min

flow rate

0.3 mL/min

pressure

3900 psi (269 bar)

column temp.

35 °C

detector

ESI-MS (+)

detector

D-erythro-Sphingosine 300.3/282.3  MRM(+) ; L-threo-Sphingosine 300.3/282.3 MRM(+) ; D-erythro-Dihydrosphingosine 302.3/284.3 MRM(+)

injection

1 μL

sample

10 ng/mL in 50% methanol/0.1% formic acid