Analysis of sphingosine 1-phosphate by capillary electrophoresis coupled to laser-induced fluorescence detection: use of a transparent fused-silica capillary.
C18-sphingosine 1-phosphate (C18-SPP) is a sphingolipid with important functional and structural roles in cells. In this paper we report a new capillary electrophoresis technique that is coupled to a laser-induced fluorescence detection (CE-LIF) method to quantify the level of C18-SPP in biological samples. The method utilizes a commercial standard C17-sphingosine 1-phosphate (C17-SPP) derivatized with naphthalene-2,3-dicarboxaldehyde (NDA), a fluorogenic dye used to label primary amines. The detection limit for the C17-SPP NDA was 0.54 fmol. We quantified the C18-SPP in leukemic human cells before and after irradiation by gamma rays. We demonstrated that the amounts of this sphingolipid decreased after the irradiation. In a second part of this work, we used the technique to evaluate the ability of a novel transparent fused-silica capillary, which allows the use of fused-silica capillaries without burning a window. Solarization, homogeneity, and sensitivity were studied using this capillary. The results demonstrate that this new durable capillary can provide a sensitive and reproducible quantitation procedure for CE-LIF studies.
Université Paul Sabatier, IMRCP, Toulouse, France.
Georges
G
Rammouz
Julnar
J
Usta
Sidney J L
SJ
Ribeiro
Younes
Y
Messaddeq
François
F
Couderc
Véréna
V
Poinsot
D000179:Acrylates / Q000737:chemistry; D045744:Cell Line, Tumor; D019075:Electrophoresis, Capillary / Q000295:instrumentation* / Q000379:methods*; D006801:Humans; D007094:Imides / Q000737:chemistry; D007700:Kinetics; D007834:Lasers; D008246:Lysophospholipids / Q000032:analysis* / Q000737:chemistry; D015394:Molecular Structure; D012822:Silicon Dioxide / Q000737:chemistry*; D013050:Spectrometry, Fluorescence / Q000295:instrumentation* / Q000379:methods*; D021241:Spectrometry, Mass, Electrospray Ionization; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000032:analysis / Q000737:chemistry; D053719:Tandem Mass Spectrometry
D000179:Acrylates; D007094:Imides; D008246:Lysophospholipids; C060506:sphingosine 1-phosphate; D012822:Silicon Dioxide; C036658:acrylic acid; D013110:Sphingosine