The supramolecular organization of Doxorubicin (DOX) within the standard Doxoves (R) liposomal formulation (DOX (R)) is investigated using visible light and phasor approach to fluorescence lifetime imaging (phasor-FLIM). First, the phasor-FLIM signature of DOX (R) is resolved into the contribution of three co-existing fluorescent species, each with its characteristic mono-exponential lifetime, namely: crystallized DOX (DOXc, 0.2 ns), free DOX (DOXf, 1.0 ns), and DOX bound to the liposomal membrane (DOXb, 4.5 ns). Then, the exact molar fractions of the three species are determined by combining phasor-FLIM with quantitative absorption/fluorescence spectroscopy on DOXc, DOXf, and DOXb pure standards. The final picture on DOX (R) comprises most of the drug in the crystallized form (similar to 98%), with the remaining fractions divided between free (similar to 1.4%) and membrane-bound drug (similar to 0.7%). Finally, phasor-FLIM in the presence of a DOX dynamic quencher allows us to suggest that DOXf is both encapsulated and non-encapsulated, and that DOXb is present on both liposome-membrane leaflets. We argue that the present experimental protocol can be applied to the investigation of the supramolecular organization of encapsulated luminescent drugs/molecules all the way from the production phase to their state within living matter.

Fluorescence lifetime microscopy unveils the supramolecular organization of liposomal Doxorubicin / Tentori, P.; Signore, G.; Camposeo, A.; Carretta, A.; Ferri, G.; Pingue, P.; Luin, S.; Pozzi, D.; Gratton, E.; Beltram, F.; Caracciolo, G.; Cardarelli, F.. - In: NANOSCALE. - ISSN 2040-3372. - 14:25(2022), pp. 8901-8905. [10.1039/d2nr00311b]

Fluorescence lifetime microscopy unveils the supramolecular organization of liposomal Doxorubicin

Pozzi D.;Caracciolo G.;
2022

Abstract

The supramolecular organization of Doxorubicin (DOX) within the standard Doxoves (R) liposomal formulation (DOX (R)) is investigated using visible light and phasor approach to fluorescence lifetime imaging (phasor-FLIM). First, the phasor-FLIM signature of DOX (R) is resolved into the contribution of three co-existing fluorescent species, each with its characteristic mono-exponential lifetime, namely: crystallized DOX (DOXc, 0.2 ns), free DOX (DOXf, 1.0 ns), and DOX bound to the liposomal membrane (DOXb, 4.5 ns). Then, the exact molar fractions of the three species are determined by combining phasor-FLIM with quantitative absorption/fluorescence spectroscopy on DOXc, DOXf, and DOXb pure standards. The final picture on DOX (R) comprises most of the drug in the crystallized form (similar to 98%), with the remaining fractions divided between free (similar to 1.4%) and membrane-bound drug (similar to 0.7%). Finally, phasor-FLIM in the presence of a DOX dynamic quencher allows us to suggest that DOXf is both encapsulated and non-encapsulated, and that DOXb is present on both liposome-membrane leaflets. We argue that the present experimental protocol can be applied to the investigation of the supramolecular organization of encapsulated luminescent drugs/molecules all the way from the production phase to their state within living matter.
2022
fluorescence lifetime; microscopy; doxorubicin
01 Pubblicazione su rivista::01a Articolo in rivista
Fluorescence lifetime microscopy unveils the supramolecular organization of liposomal Doxorubicin / Tentori, P.; Signore, G.; Camposeo, A.; Carretta, A.; Ferri, G.; Pingue, P.; Luin, S.; Pozzi, D.; Gratton, E.; Beltram, F.; Caracciolo, G.; Cardarelli, F.. - In: NANOSCALE. - ISSN 2040-3372. - 14:25(2022), pp. 8901-8905. [10.1039/d2nr00311b]
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11573/1706108
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