1 research outputs found
Validated Method for the Quantification of Free and Total Carnitine, Butyrobetaine, and Acylcarnitines in Biological Samples
A validated
quantitative method for the determination of free and
total carnitine, butyrobetaine, and acylcarnitines is presented. The
versatile method has four components: (1) isolation using strong cation-exchange
solid-phase extraction, (2) derivatization with pentafluorophenacyl
trifluoromethanesulfonate, (3) sequential ion-exchange/reversed-phase
(ultra) high-performance liquid chromatography [(U)ÂHPLC] using a strong
cation-exchange trap in series with a fused-core HPLC column, and
(4) detection with electrospray ionization multiple reaction monitoring
(MRM) mass spectrometry (MS). Standardized carnitine along with 65
synthesized, standardized acylcarnitines (including short-chain, medium-chain,
long-chain, dicarboxylic, hydroxylated, and unsaturated acyl moieties)
were used to construct multiple-point calibration curves, resulting
in accurate and precise quantification. Separation of the 65 acylcarnitines
was accomplished in a single chromatogram in as little as 14 min.
Validation studies were performed showing a high level of accuracy,
precision, and reproducibility. The method provides capabilities unavailable
by tandem MS procedures, making it an ideal approach for confirmation
of newborn screening results and for clinical and basic research projects,
including treatment protocol studies, acylcarnitine biomarker studies,
and metabolite studies using plasma, urine, tissue, or other sample
matrixes