Direct determination of thiolated amino acids using poly(dimethylsiloxane) microchip capillary electrophoresis coupled with electrochemical detection
The determination of the biomarkers homocysteine, glutathione and N-acetyl-L-cysteine has been investigated using microchip capillary electrophoresis coupled with pulsed amperometric detection (PAD). The microchip configuration consists of a first layer of polydimethylsiloxane (PDMS) composed of injection and separation channels, reservoirs and a gold microwire sealed with a second layer of PDMS. A gold microwire was used as the working electrode and platinum microwire
located in the waste reservoir was used as a counter electrode. The experiments were carried out with 20 mM boric acid buffer (pH 8.5) and compared to 20 mM MES and 1 mM SDS buffer (pH 6.0) at the detection potential of 0.7 V. The effect of including injection and separation potential, pH and injection time, and PAD parameters were studied to optimize the separation and detection. The results showed that homocysteine, glutathione and N-acetyl-L-cysteine could be separated and detected in less than 100 s with 20 mM MES and 1 mM SDS buffer (pH 6.0), with more reproducibility and sensitivity than a standard boric acid buffer. This method was applied to determine homocysteine, glutathione and N-acetyl-L-cysteine in human plasma, with the intent indicating cardiovascular disease.