Instrumental innovations in capillary electrophoresis with electrochemical detection in the analysis of mono and oligosaccharides, amino acids and proteins / Inovações instrumentais em sistemas de eletroforese capilar com detecção eletroquímica e aplicações em análises de mono e oligossacarídeos, aminoácidos e proteínas

AUTOR(ES)
DATA DE PUBLICAÇÃO

2008

RESUMO

This work shows the development of two equipments (H1 and B1) of capillary electrophoresis (CE) with contactless conductivity detection (C4D) applied to the analysis of biomolecules. They have a system named MSE (module for separated electrolysis) that avoids the harmful effect of electrolysis. H1 have only one capillary and the majority of the experiments presented here were developed in this equipment. It also have a system of thermal marks (TM) used to correct the EOF effect on the migration of ions Na+ e K+ in egg white. We also developed the separation and detection of proteins (10 µmol·L-1) between 12 an 66 kDa, showing that C4D can be used to detect these molecules using substances to avoid adsorption on the capillary wall. Experiments of separation and detection of chitooligosaccharides enzymatically produced were also developed in H1. By using NaOH and acetonitrile as the electrolyte, we did the complete separation of six chitooligosaccharides (C1 to C6) with limits of detection and quantification less than 3 µmol·L-1 and 10 µmol·L-1 , respectively. After the enzymatic assays of C2 to C6 with the chitinase purified from the beetle Tenebrio molitor (TmChi), it is observed that this enzyme cut these substrates with very low efficiency, as expected. This enzyme also cut C4 producing C2 and cut C5 producing C2 and C3. C5 is the best substrate for this enzyme. C6 produces C2 and C3, showing that this enzyme is a endo- chitinase type. The equipment B1 has eight capillaries and eight C4D detectors with the best signal/noise ratio at 1 MHz e 4 Vpeak-to-peak . By using B1, it is possible run up to eight different samples with four different electrolytes and separation potentials. In this equipment, we develop the separation of 20 proteinogenic amino acids (AAs) using two different separation conditions at low pH. Separations of these molecules using high-pH electrolytes and with different potentials were also demonstrated. The development of a microchip of PDMS with an immobilized enzyme reactor (IMER) to the glucose detection was also constructed. The detection of hydrogen peroxide produced by the enzyme glucose oxidase linked on the IMER was measured by amperometry. The performance of this chip was evaluated with glucose and peroxide injections. The best potential for the oxidation of the hydrogen peroxide was 0.9V, using electrolyte at pH 8.5 and 1100 V as the potential of separation. A linear curve was observed between peak current and glucose concentration in the range from 0.1 up to 6.2 mmol·L-1 . Determinations in soda shows 216 mmol·L-1 of glucoce, that is a good agreement with other reports.

ASSUNTO(S)

lab-on-a-chip lab-on-a-chip eletroforese capilar detecção condutométrica sem contato detection of biomolecules contacteless conductivity detection capillary electrophoresis detecção de biomoléculas

ACESSO AO ARTIGO

Documentos Relacionados