QUANTIFICATION OF FREE SIALIC ACID IN URINE AND CEREBROSPINAL FLUID BY LC–TANDEM MASS SPECTROMETRY: ONE METHOD FOR TWO DIFFERENT MATRICES

Background: Disturbance in sialic acid (SA) metabolism has been described in genetic disorders that involve its transport (Salla disease) or its overproduction (sialuria). A change in its metabolism has also been described in many non-genetic pathological conditions (e.g. infections, inflammations, endocrinological, cardiovascular, neurological, and renal pathologies).Analysis of free sialic acid (FSA) in urine and cerebrospinal fluid (CSF) is a very important tool for diagnosing and distinguishing these conditions. In human CSF and urine, the determination of FSA has been carried out by several methods. Some of those methods are either relatively insensitive and lack specificity or require relatively high CSF volume and can never be applied in routine diagnostic setting. Liquid chromatography–tandem mass spectrometry (LC/MS/MS) has the advantage of improved sensitivity, selectivity and the advantage of ease of operation compared with the more elaborate procedures. In this study, we describe LC/MS/MS method that is robust enough to simultaneously measure FSA in the two significantly different matrices of urine and CSF.

Methods: CSF or urine sample were deproteinized by acetonitrile and subjected to LC-MS/MS analysis using negative electrospray ionization (ESI). Isotope dilution method was used for quantification of FSA.

Results: Limit of detection (LOD) of our method was 0.50μM in both CSF and urine. Intra- and inter-assay variations of CSF analysis were 4.54% and 14.1%; respectively. Those of urine analysis were 4.77% and 12.4%; respectively.

Conclusions: This validated method allows simultaneous FSA determination in urine and CSF rapidly with high sensitivity, specificity and robustness.