Fabry disease is an X linked hereditary metabolic disease that causes renal failure and many lifethreatening complications as a result of alphagalactosidase deficiency.
The enzymatic deficiency causes to the lysosomal accumulation of globotriaosylceramide (Gb3), and the deacylated derivative of Gb3, globotriaosylsphingosine (LysoGb3). Therefore, the measurement of plasma LysoGb3 has become an important biomarker for diagnosis and as a therapeutic monitoring of enzyme replacement therapy. In this study we aimed to develop a simple, fast, cheap, underivatized LC MS MS method for measurement of plasma LysoGB3 levels. The chromatographic
separation was operated on an Acquity UPLC C18 (50 mm × 2.1 mm, 1.7 μm) with gradient elution using %98 %2 Water/Methanol containing 0.05% formic acid and methanol as the mobile phase at the flow rate of 0.4 ml/min and total run time was only 3 minutes. The mass quantification was carried on the multiple reaction
monitoring (MRM) of the transitions of m/z 786.2 → 282.1 for LysoGB3 and m/z 624.2 → 282.1 for LysoGB2 (the internal standard), respectively. First of all we
investigated the effect of liquidliquid extraction solvents in terms of peak shape, peak response, matrix effect and recovery. In the scope of method validation parameters, selectivity, linearity, determination and quantitation limits, carryover effect, reproducibility, recovery, matrix effect, reference range verification and stability
studies were carried out. The limit of detection was found as 0.0018 ng/ml and the limit of quantification was found as 0.0053 ng/ml. The intraday and interday
reproducibility studies are below 7% which is suitable for validation criteria. Recovery rate was found as 96102 % while matrix effect was found as 98103%. At last,the validated method was successfully performed in 12 Fabry patients and 80 healthy individuals.