S.S. Espinoza et al.
Journal of Photochemistry & Photobiology A: Chemistry 364 (2018) 140–144
to irradiate samples for cycles of 5 min at 150 mV. Each of one consisted
of the use of flashing light 0.5 s on/ 0.5 s off.
The ROS production was explored through the reaction between 4-
aminoantipyrine and phenol in the presence of Lipo-D-ODA yielding the
corresponding quinoneimine [14]. An aliquot of 2 mL of Lipo-D-ODA
solution (dilution ratio 1:4 in phosphate buffer 30 mM pH 7.00) was
added to 4 mL of a mixture (v/v) of phenol (0.1 mol/L), 4-aminoanti-
−3
pyrine (1.5 × 10
mol/L) and the phosphate buffer at pH 7.0
(
0.10 M). The photocatalytic reaction started upon direct irradiation of
laser light of 405 nm. The solution was collected and centrifuged, and
the supernatant was measured at 505 nm. This procedure was carried
out at regular intervals for about 20 min.
3. Result and discussion
A photosensitive material was obtained by the incorporation of a
highly hydrophobic porphyrin, D-ODA, into liposomes (Lipo-D-ODA)
through self-assembling of the aliphatic chains. Porphyrins are second
generation sensitizers [15], producing reactive oxygen species (ROS) by
2 2
Fenton reaction (H O and porphyrins) or by light (porphyrins).
Considering that significant spectral changes are observed upon
transfer of a porphyrin molecule from an aqueous phase to the lipid
phase, we evaluate the spectroscopic response of D-ODA and deuter-
oporphyrin IX in the liposome system. Fig. 1A corresponds to the ab-
sorbance at 405 nm of the two compounds; the curves are identical,
proving that both porphyrins reach the same concentration in the or-
ganic medium. On the other hand, the fluorescence spectrum shows a
different behavior. Whereas the emission intensity of deuteroporphyrin
IX decreases with increasing concentration, due to aggregation-caused
quenching (ACQ) [16], the D-ODA response follows a hyperbolic-type
curve (Fig. 1B). Clearly, the hydrophobic interactions with the lipidic
environment destabilize the aggregated species in favor of the in-
corporation of the monomer into the liposome, resulting in the sig-
nificant reduction of the quenching induced by aggregation.
To evaluate the efficiency of production of ROS, we measured the
absorption of the chromophore quinoneimine at 505 nm, which is in-
dicative of the efficiency of the reaction of the phenolic substrate and 4-
aminoantipyrine. This reaction involves ROS produced in the presence
of photoexcited porphyrins. Fig. 2 shows the absorption at 505 nm vs
time of irradiation at 405 nm; two different curves are obtained in the
absence (b) or presence(c) of 50 μl hydrogen peroxide (50 mM). In both
cases, the excited Lipo-D*-ODA could activate the ground state mole-
cular oxygen to produce oxygen singlet or superoxide anion radical
according to type II and I mechanism respectively [17]. It is interesting
to notice that higher production of ROS is observed in (c), due to the
Fig. 3. Cyclic voltammograms obtained during the evaluation of the release of
(A) Quercetin (12.6 ppm) and (B) Fc-ODA (21 ppm) in different liposome sys-
tems with or without irradiation at 405 nm, in 10 mM PBS buffer, 0.1 M NaCl,
−
1
pH 7.20. Scan rate 50 mV s
.
4. Conclusions
A new photosensitive liposome, containing a highly hydrophobic
porphyrin as sensitizer was obtained. This new material resulted in an
efficient nanocarrier of hydrophobic analytes that were delivered by
light stimuli in aqueous solution and further electrochemically ana-
lyzed. Considering that biomacromolecules suit well in the lipidic en-
vironment, it can be envisioned for future research including membrane
proteins, ADN and other macromolecules.
2 2
formation of reactive species from H O (Fenton reaction). Meanwhile,
the curve (a) demonstrates non-production of ROS without irradiation.
To establish a strategy of measurement in an aqueous medium of
hydrophobic compounds electrochemically active, we evaluated the
analyte release by light excitation of the D-ODA liposome with the light
of 405 nm. In Fig. 3, the cyclic voltammetries show different response
depending on the structure of the electrochemically active compound,
in 10 mM PBS buffer, 0.1 M NaCl, pH 7.20. The quercetin oxidation at
Acknowledgements
Financial support from the University of Buenos Aires (UBACyT
2014-201720020130100469BA), ANPCyT (PICT 2013-1541) and
CONICET (11220130100029CO) are gratefully acknowledged. S.S.E.
would like to thank CONICET for her postdoctoral fellowship.
200 mV was only observed after irradiation on the Lipo-D-ODA-Quer-
cetin, indicating that it is embedded in the liposome and requires
membrane rupture. On the contrary, Fc-ODA displays an electro-
chemical signal before the treatment with light, which can be explained
considering that the hydrophobic portion (ODA) is embedded in the
membrane, and the polar moieties (Fc) oriented to the outer, aqueous
space. Furthermore, the ferrocene signal increases after irradiation
probably due to the exposition of more redox groups. However, the
anodic potential shifts to a more positive value indicating the passiva-
tion effect of the aliphatic matrix.
Appendix A. Supplementary data
References
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Espinoza, Silvia Soto
