Nona Semana Integrada do Instituto de Física de São Carlos

Optical characterization of metabolics fluorophores NADH and FAD in solution

Não agendado

20m

Iniciação Científica

Palestrante

Giancarlo Souza

Descrição

Cell metabolism is the set of biochemical processes that occurs in an organism to energy production, cellular development and maintain cell vital functions. (1) Monitoring cellular metabolism can be used to distinguish between healthy and abnormal cells. It can be done by observing some cellular organelles that have an important role in cellular metabolism. In this context, mitochondrial activity can be monitored by native fluorescent electron carriers called NADH and FAD. (2) The optical redox ratio can be calculated between these two coenzymes can be used to estimate cell metabolic process. Therefore, optical characterization of these and other biomolecules is essential to the monitoring cellular metabolism. (3) In this study, synthetic NADH and FAD solutions (Sygma Aldrich) were prepared at different concentrations and, interactions between fluorophores and medium were evaluated by fluorescence spectroscopy. Measurements of stationary state fluorescence and fluorescence lifetime spectroscopy were performed to characterize these biomolecules in solution. For steady-state fluorescence comparison between the two fluorophores, an excitation-emission matrix (Ex. 250-500 nm, Em. 300-650 nm) was determine up for each one of the solutions. Fluorescence lifetime was measured by TCSPC, using a 20 MHz pulsed laser emitting at 378 nm and processed in MATLAB. Average fluorescence lifetimes were calculated also for comparison. Results indicates average lifetime of NADH decreased from 2.10 ns to 1.10 ns when comparing 20 uM NADH solution with 20 uM NADH and 20 uM FAD solution. While excitation-emission matrix showed a possible energy transfer between the fluorophores when concentrations achieved 20 uM NADH and 0.2 uM FAD (Figure 1 (A) and (B) ), matrices shows a decrease of intensity of NADH fluorescence, when FAD is present in the solution.

Referências

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