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Simultaneous determination of 18 D-amino acids in rat plasma by an ultrahigh-performance liquid chromatography-tandem mass spectrometry method: application to explore the potential relationship between Alzheimer’s disease and D-amino acid level alterations

Xing, Yuping, Li, Xiaoyan, Guo, Xingjie, Cui, Yan
Analytical and bioanalytical chemistry 2016 v.408 no.1 pp. 141-150
Alzheimer disease, acetonitrile, biomarkers, derivatization, hydrogen, ionization, liquid chromatography, monitoring, pathogenesis, phenylalanine, rats, tandem mass spectrometry
D-Amino acids are increasingly being recognized as important signaling molecules, and abnormal levels of D-amino acids have been implicated in the pathogenesis of Alzheimer’s disease. To evaluate the potential relationship between Alzheimer’s disease and D-amino acids, a simple, sensitive, and reliable UPLC-MS/MS method with pre-column derivatization was developed and validated for simultaneous determination of 18 D-amino acids in rat plasma. The analytes were extracted from plasma samples by a protein precipitation procedure, and then derivatized with (S)-N-(4-nitrophenoxycarbonyl) phenylalanine methoxyethyl ester [(S)-NIFE]. Chromatographic separation was achieved using an ACQUITY UPLC BEH C₁₈ column (2.1 mm × 50 mm, 1.7 μm) with a mobile phase composed of acetonitrile containing 8 mM ammonium hydrogen carbonate at a flow rate of 0.6 mL min⁻¹. The analytes were detected by electrospray ionization in positive ion multiple reaction monitoring modes. Under the optimum experimental conditions, all the linear regressions were acquired with r > 0.9932. The limits of quantitation of all derivatized D-amino acids were within 0.05–40.0 ng mL⁻¹ in rat plasma. The intra- and inter-day precisions, expressed as percentage relative standard deviations (%RSD), were within the range of 12.3 and 10.1 %, respectively. The recoveries for all the analytes were observed over the range of 82.8–100.5 % with RSD values less than 12.5 %. Finally, the proposed method was successfully applied to simultaneous determination of the 18 D-amino acids in plasma from Alzheimer’s disease rats and age-matched normal controls. Results showed that the concentrations of D-serine, D-aspartate, D-alanine, D-leucine, and D-proline in Alzheimer’s disease rat plasma were significantly decreased compared with those in normal controls, while D-phenylalanine levels increased. It was revealed that some of these D-amino acids would be potential diagnostic biomarkers for Alzheimer’s disease. Graphical abstract Simultaneous determination of 18 D-amino acids in rat plasma.