Development of capillary electrophoresis with capacitively coupled contactless conductivity detection for clinical analysis and related applications

This thesis was focused on the development of capillary electrophoresis with
capacitively coupled contactless conductivity detection (CE-C4D) for quantification
of small organic ions that are poorly UV absorbing molecules in clinical analysis and
related applications. CE-C4D is suitable for clinical analysis that usually involves
small sample volumes in the nanolitre range. Due to the high separation efficiency
obtained measuring the analyte ions in the presence of complex matrices is possible.
The determination of uric acid was firstly examined to explore our knowledge for the
routine compound analysis. We successfully determined uric acid in clinical samples
in its anionic form using MES/His buffer. The results obtained were well correlated
with the standard enzymatic method. We further investigated the determination of
non-UV absorbing molecules, namely lactate, carnitine and acylcarnitines. CE-C4D
proved to be suitable for the analysis of these compounds with little sample
preparation and without any derivatization steps. Plasma lactate, which is an
important parameter to evaluate the anaerobic metabolism and the exercise intensity
in terms of the lactate threshold, was quantified. We successfully performed the
quantification of plasma lactate in its anionic form using MES/His buffer. The
plasma lactate concentrations at different exercise conditions and the lactate threshold
curve were achieved. The results obtained by CE-C4D corresponded to the results
observed by the enzymatic method. We also demonstrated the determination of
carnitine and its derivatives (acylcarnitines) in various clinical samples. Free
carnitine, short- and medium-chain acylcarnitines, consisting of acetyl-, propionyl-,
isovaleryl-, hexanoyl-, octanoyl-, and valproyl-carnitines in plasma and urine samples
were successfully quantified. The similarity of their chemical structures posed a high
challenge in this study and required comprehensive optimization of the running
buffer. An acetic acid buffer at pH 2.6 in the presence of hydroxypropyl-β-
cyclodextrin (HP-β-CD) as a modifier was found to be the optimum. The isomers of
valproyl- and octanoyl-carnitines could additionally be distinguished. The results
obtained by CE-C4D were compared to standard enzymatic- and LC/MS-methods. In
food analysis, the determination of lactic acid and carnitine in various food samples
were also achieved. The determination of D- and L-lactic acid in fresh and spoiled
milk and yogurt samples were successfully obtained in the presence of Tris/Maleic
acid buffer using vancomycin as a chiral selector. In addition, the determination of
carnitine in food samples was investigated in the presence of acetic acid buffer at pH
2.6. The carnitine contents in various samples were successfully quantified with good
method reproducibility, and the results obtained were acceptable.