Analysis of chiral amino acids in cerebrospinal fluid samples linked to different stages of Alzheimer disease

Chiral micellar electrokinetic chromatography with laser-induced fluorescence detection (chiral-MEKC-LIF) was used to investigate D- and L-amino acid contents in cerebrospinal fluid (CSF) samples related to different Alzheimer disease (AD) stages. CSF samples were taken from (i) control subjects (S1 pool), (ii) subjects showing a mild cognitive impairment who remained stable (S2 pool), (iii) subjects showing an mild cognitive impairment that progressed to AD (S3 pool) and (iv) subjects diagnosed with AD (S4 pool). The optimized procedure only needed 10 μL of CSF and it included sample cleaning, derivatization with FITC and chiral-MEKC-LIF separation. Eighteen standard amino acids were baseline separated with efficiencies up to 703,000 plates/m, high sensitivity (LODs in the nM range) and good resolution (values ranging from 2.6 to 9.5). Using this method, L-Arg, L-Leu, L-Gln, γ-aminobutyric acid, L-Ser, D-Ser, L-Ala, Gly, L-Lys, L-Glu and L-Asp were detected in all the CSF samples. S3 and S4 samples (i.e. AD subjects) showed significant lower amounts of L-Arg L-Lys, L-Glu and L-Asp compared to the non-AD S1 and S2 samples, showing in the S4 group the lowest amounts of L-Arg L-Lys, L-Glu and L-Asp. Moreover, γ-aminobutyric acid was significantly higher in AD subjects with the highest amount also found for S4. No significant differences were observed for the rest of amino acids including D-Ser. Based on the obtained chiral-MEKC-LIF data, it was possible to correctly classify all the samples into the four groups. These results demonstrate that the use of enantioselective procedures as the one developed in this work can provide some new light on the investigations of AD, including the discovery of new biomarkers related to different stages of AD.