RESUMO
Three alkylated DNA adducts, N3methyladenine, N3ethyladenine and N7ethylguanine, have been proved to be potential biomarkers for DNA injury caused by exposure to cigarette smoke. In this study, a highly specific and sensitive method using a new mixed-mode sulfonate-functionalized poly(glycidyl methacrylate-divinylbenzene) as a solid-phase extraction sorbent was developed for the analysis of these three alkylated-purine adducts in human urine. Under optimized conditions, the prepared sorbent interacts strongly with these urinary adducts, demonstrating high clean-up efficiency and extraction recovery. The method detection limits (S/Nâ¯≥â¯3) of N3-MeA, N3-EtA and N7-EtG were 1.75, 0.20, and 0.15â¯pgâ¯mL-1, respectively, while the method quantitation limits were found to be 5.78, 0.66, and 0.49â¯pgâ¯mL-1 for N3-MeA, N3-EtA and N7-EtG, respectively. The intra-day and inter-day precisions were investigated, of which were in the range of 1.6-3.8% and 3.2-5.6%, respectively. The recovery values of the alkylated DNA adducts in spiked urine sample were ranged 89.7-104.5%. Their concentrations were statistically significantly higher in smokers than in nonsmokers. These results show that the proposed method is suitable for the analysis of alkylated DNA adducts.
Assuntos
Cromatografia Líquida/métodos , Adutos de DNA/urina , Microesferas , Purinas/urina , Espectrometria de Massas em Tandem/métodos , Adulto , Alquilação , Biomarcadores , Adutos de DNA/química , Adutos de DNA/isolamento & purificação , Compostos de Epóxi/química , Humanos , Modelos Lineares , Masculino , Metacrilatos/química , Purinas/química , Purinas/isolamento & purificação , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Fumar , Extração em Fase Sólida/métodos , Ácidos Sulfônicos/química , Compostos de Vinila/química , Adulto JovemRESUMO
The interaction of DNA with Bis [N,N-bis (carboxymethyl) aminomethyl] fluorescein-Ferrous(III) (Fluorescein-DA-Fe(III)) with dual functional (sonodynamic and sonocatalytic) activity was studied by UV-vis spectroscopy, fluorescence spectroscopy, FT-IR spectroscopy, circular dichroism (CD) spectroscopy and viscosity measurements. And then, the damage of DNA caused by Fluorescein-DA-Fe(III) under ultrasonic irradiation (US) was researched by agarose gel electrophoresis and cytotoxicity assay. Meanwhile, some influenced factors such as ultrasonic irradiation time and Fluorescein-DA-Fe(III) concentration on the damage degree of DNA molecules were also examined. As a control, for Bis [N,N-bis (carboxymethyl) aminomethyl] fluorescein (Fluorescein-DA), the same experiments were carried out. The results showed that both Fluorescein-DA-Fe(III) and Fluorescein-DA can interact with DNA molecules. Under ultrasonic irradiation, Fluorescein-DA shows sonodynamic activity, which can damage DNA molecules. While, in the presence of Fe(III) ion, the Fluorescein-DA-Fe(III) displays not only sonodynamic activity but also sonocatalytic activity under ultrasonic irradiation, which injures DNA more serious than Fluorescein-DA. The researches confirmed the dual function (sonodynamic activity and sonocatalytic activity) of Fluorescein-DA-Fe(III) and expanded the usage of Fluorescein-DA-Fe(III) as a sonosensitizer in sonodynamic therapy (SDT).