Estudos de membranas modelo e efeitos de terpenos em membranas de leishmania por ressonância paramagnética eletrônica
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Data
2013-11-16
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Universidade Federal de Goiás
Resumo
Electron paramagnetic resonance (EPR) spectroscopy of spin labels was used to
study the main structural accommodations of environment-sensitive probes in the bilayers of
saturated phosphatidylcholines with acyl chains lengths ranging from 16 to 22 carbon atoms.
The more detailed analysis were made on the spin probe 5-doxyl methyl stearate (5-DMS)
whose EPR spectra allowed to identify two distinct spectral components in thermodynamic
equilibrium at temperatures below and above the main phase transition. The EPR spectroscopy
distinguishes two components associating lower motion with higher polarity (denoted
component 1) and higher motion with lower polarity (component 2), which may be assigned
to one shallow (more rigid structure) and one deep population of spin probe, respectively.
At temperatures until 22◦C only one spectral component can be noted in the spectra whereas
at 30◦C the component 1 coexists with an appreciable fraction of component 2. In the
liquid-crystalline phase the 5-MSL showed two spectral components for all studied lipids in
the entire range of measured temperatures. An accurate analysis of EPR spectra, performed
using two fitting programs (NLLS and EPRSIM), allowed us to obtain the thermodynamic
profile to these major probe accommodations. Focusing the analysis on two-component EPR
spectra, it was studied the influences of cholesterol and a membrane permeation enhancer
on the mobility and distribution of spin label on these two main bilayer environments.
Parte II
Cutaneous leishmaniasis is a neglected tropical disease that infects millions of
people worldwide, representing a serious public health problem. The current treatment is
based on chemotherapy, using pentavalent antimonials compounds, which cause serious side
effects. Electron paramagnetic resonance (EPR) spectroscopy of the spin-label analog of
stearic acid (5-DSA ) was used to monitor the effect of the terpenes α-terpineol, 1,8-cineole,
III
(+)-limonene and nerolidol on the plasma membrane fluidity of Leishmania amazonensis
promastigotes. Cytotoxic effects on the parasite were also measured to investigate the relationship
between the cytotoxic potential of terpenes and their ability to alter membrane
fluidity. All terpenes increased the fluidity of the cellular membrane, without significant differences
at higher concentrations. However, the minimum concentration required to cause a
change in the membrane was very different between the terpenes and similar to that caused
50% growth inhibition (IC50) showing a correlation between membrane alterations and cytotoxicity.
The IC50 values of terpenes analyzed showed the following relationship: nerolidol
< (+)-limonene < α-terpineol < 1,8-cineole, with an IC50 of 8 μM for nerolidol and 4700 μM
to 1,8- cineole. The EPR spectra of the maleimide derivative spin label (6-MSL) covalently
attached to the Leishmania membrane proteins indicated that the terpenes essentially do not
alter the dynamics of protein backbone and only increase the mobility of the nitroxide side
chain. Cell lysis was not detected at cytotoxic concentrations, as measured by the presence
of spin-labeled membrane fragments. Since the terpenes are considered potent skin permeation
enhancers with low irritation potential, this work suggests checking the possibility of
terpenes applications in the treatment of tegumentary leishmaniasis, where terpenes could
perhaps perform a dual action of be an active principle and at the same time facilitate the
penetration of other molecules with antileishmanial activity.
Descrição
Palavras-chave
Componentes espectrais , RPE , Terpenos , Leishmania , Spectral components , EPR , Terpenes , Leishmania
Citação
CAMARGOS, Heverton Silva de. Estudos de membranas modelo e efeitos de terpenos em membranas de leishmania por ressonância paramagnética eletrônica. 2013. 169 f. Tese (Doutorado em Física) - Universidade Federal de Goiás, Goiânia, 2013.