High-performance liquid chromatographic separation of acylcarnitines following derivatization with 4'-bromophenacyl trifluoromethanesulfonate.
A high-performance liquid chromatographic method for the separation of acylcarnitines after derivatization with 4'-bromophenacyl trifluoromethanesulfonate is presented. Derivatization of acylcarnitines was achieved at room temperature within 10 min. Separation of the acylcarnitine 4'-bromophenacyl esters was accomplished by high-performance liquid chromatography using as the analytical column a Resolve-PAK 5-microns C18 radially compressed cartridge eluted with a tertiary gradient containing varying proportions of water, acetonitrile, tetrahydrofuran, triethylamine, potassium phosphate, and phosphoric acid. Acylcarnitine 4'-bromophenacyl esters were detected spectrophotometrically at 254 nm. Baseline separation was obtained for a standard mixture (5 nmol of each injected) containing carnitine, acetyl-, propionyl-, butyryl-, valeryl-, hexanoyl-, heptanoyl-, octanoyl-, nonanoyl-, decanoyl-, lauroyl-, myristroyl-, palmitoyl-, and stearoylcarnitine. Nearly complete separation was obtained for a standard mixture containing butyryl-, isobutyryl-, isovaleryl-, and 2-methylbutyrylcarnitine. The method was applied to a normal human urine and then to this same urine spiked with the acylcarnitine standards. Urinary acylcarnitine profiles from patients having propionic acidemia, isovaleric acidemia, and medium-chain acyl-CoA dehydrogenase deficiency were performed. Urinary isovalerylcarnitine was quantified in the patient with isovaleric acidemia using heptanoylcarnitine as an internal standard.
Minkler
P E
PE
Medical Research, Veterans Administration Medical Center, Cleveland, Ohio 44106.
Ingalls
S T
ST
Hoppel
C L
CL
D000098:Acetophenones; D000108:Acetylcarnitine / Q000652:urine*; D042964:Acyl-CoA Dehydrogenase; D044944:Acyl-CoA Dehydrogenases / Q000172:deficiency; D002250:Carbon Radioisotopes; D002331:Carnitine / Q000031:analogs & derivatives* / Q000627:therapeutic use / Q000652:urine; D055598:Chemical Phenomena; D002621:Chemistry; D002851:Chromatography, High Pressure Liquid; D006801:Humans
D000098:Acetophenones; D002250:Carbon Radioisotopes; C027333:3-methylbutyrylcarnitine; D000108:Acetylcarnitine; C043597:4'-bromophenacyl triflate; D044944:Acyl-CoA Dehydrogenases; D042964:Acyl-CoA Dehydrogenase; D002331:Carnitine