Neurológiai betegségek (pl. Alzheimer-kór, Huntington-kór, Parkinson-kór)
The simultaneous quantitative estimation of tryptophan (TRP) and its metabolites represents
a great challenge because of their diverse chemical properties, e.g., presence of
acidic, basic, and nonpolar functional groups and their immensely different concentrations
in biological matrices. A short ultra high-performance liquid chromatography (UHPLC)-tandem
mass spectrometry (MS/MS) method was validated for targeted analysis of TRP and its
11 most important metabolites derived via both kynurenine (KYN) and serotonin (SERO)
pathways in human serum and cerebrospinal fluid (CSF): SERO, KYN, 3-hydroxyanthranilic
acid, 5-hydroxyindoleacetic acid, anthranilic acid, kynurenic acid (KYNA), 3-hydroxykynurenine
(3-HK), xanthurenic acid, melatonin, picolinic acid (PICA), and quinolinic acid (QUIN).
After selecting the "best" reversed-phase column and organic modifier, DryLab®4 was
used to optimize the gradient time and temperature in chromatographic separation.
To achieve absolute quantification, deuterium-labeled internal standards were used.
Among all compounds, 3 were analyzed in derivatized (butyl ester) forms (3-HK, PICA,
and QUIN) and the remaining 9 in underivatized forms. Validation was performed in
accordance with the ICH and FDA guidelines to determine the intraday and interday
precision, accuracy, sensitivity, and recovery. To demonstrate the applicability of
the developed UHPLC-MS/MS method, the aforementioned metabolites were analyzed in
serum and CSF samples from patients with multiple sclerosis (multiple sclerosis group)
and those with symptomatic or noninflammatory neurological diseases (control group).
The concentration of QUIN dramatically increased, whereas that of KYNA slightly decreased
in the multiple sclerosis group, resulting in a significantly increased QUIN/KYNA
ratio and significantly decreased PICA/QUIN ratio.
