Microssistemas eletroforéticos em materiais poliméricos de duplo canal com detecção amperométrica

Abstract

Electrophoretic microsystems (EM) are powerful tools for the separation of species of microsystems analyzes which can easily be combined with electrochemical detection (ECD) and therefore making it ideal for a method of detection. However, the influence of high voltage at the working electrode used for the separation is a problem to be overcome due to the increased signal/noise ratio and possible damage of the electrode and/or the potentiostat. Thus, it was proposed in this thesis one EM hybrid PDMS / glass configuration with dual-channel potentiostat coupled to an electrically isolated in order to minimize the influence of high potential in the separation channel and improve the separation efficiency of the species and subsequently, improve detection limits. The EM contains two separate parallel channels 200 microns and a channel separation and another reference, and each containing a platinum electrode 15 or 50 μm placed about 1 to 4 μm in the channel. An electrode served as the working electrode, positioned in the separation channel, and another electrode as reference electrode, placed in the reference channel. This configuration associated with the electrically isolated potentiostat allowed the amperometric signals were measured without any change or potential interference arising from the high voltage applied separation. Aiming to evaluate the effectiveness of the methodology proposed in this thesis, samples nitrite, tyrosine and peroxynitrite (reactive nitrogen species – RNS), hydrogen peroxide (reactive oxygen species – ROS), ascorbic acid, glutathione and cysteine were injected into the channel containing the working electrode, while simultaneously boric acid buffer pH 11 containing TTAB was injected into the reference channel containing the reference electrode. From this configuration, we obtained a significant reduction in noise level (about 0.94 pA) and a relative improvement in the resolution ratified by electropherograms, compared with using single channel configuration. The limits of detection (LOD) for the chemical species mentioned above were 0.58 μM, 0.14 μM, 0.75 μM, 0.21 μM, 0.82 μM, was not obtained for cysteine and 1.63 μM, respectively. The efficiency can also be seen by analyzing nitrite performed on samples of perfusate blood of sheeps and rats, where have been detected a concentration of 68.05 μM and 22.04 μM, respectively, by the proposed method. It was also proposed in this thesis, microfabrication and evaluation of a PMMA electrophoretic microsystem with single channel configuration coupled to a base made of the same material to fix the microchip with electrochemical detection using a carbon paste electrode. The purpose of the construction of the base was to obtain, by fixing, reproducibility of events. And the microfabrication of PMMA EM aimed the viability of its use in analysis perspective as having the lowest cost per unit made due to the use of CO2 laser for microfabrication, which has a value considerably lower, compared with photolithographic processes. The evaluation of this system was performed through the analysis standards of serotonin and acetaminophen, which proved that the microfabrication of this system showed good reproducibility and repeatability of events, making it viable processing.