Comparison of the Photodynamic Effect of Exogenous Photoprotoporphyrin and Protoporphyrin IX on PAM 212 Murine Keratinocytes¶

Abstract

A comparative study of the cellular photosensitizing properties of protoporphyrin IX (PpIX) and photoprotoporphyrin (Ppp) was carried out in the transformed murine keratinocyte cell line, PAM 212. Time-course fluorescence studies were performed to determine the rate of uptake by cells together with fluorescence microscopy. The sensitized cells were laser irradiated with a range of light doses at 635 or 670 nm to determine the phototoxicity of the two compounds and to investigate their relative fluorescence photobleaching properties. Ppp showed enhanced phototoxicity at both its optimal activation wavelength of 670 nm (eight times more phototoxic than PpIX activated at its optimal wavelength of 635 nm for the same fluence) and at 635 nm (three times more phototoxic than PpIX at the same wavelength), using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. The photobleaching rate of Ppp in cells was found to be higher using 670 nm irradiation compared with that of PpIX at 635 nm irradiation. At 635 nm, however, the photobleaching rate of Ppp was comparable to that of PpIX. The photobleaching quantum yields of the two compounds in cells were found to be similar at approximately 5 x 10(-4), with the same value confirmed at both 670 and 635 nm irradiation for Ppp. The fluorescence lifetime of Ppp in cells was measured as 5.4 ns using time-correlated single photon counting.

Anti‐Stokes fluorescence from endogenously formed protoporphyrin IX – Implications for clinical multiphoton diagnostics

“…having a cross-section (σ 2PE ) of the order of 10 −50 cm 4 s, photon, competes with a more probable one-photon transition, i.e. cross-section (σ 1PE ) of the order of 10 −15 cm 2 photon [25]. The probabilities of the one-photon ( P 1PE ) and the two-photon ( P 2PE ) excitation processes can be described as…”

Protoporphyrin IX–β‐Cyclodextrin Bimodal Conjugate: Nanosized Drug Transporter and Potent Phototoxin

“…Exogenous administration of ALA and its esters (8) practically bypasses the negative feedback control exerted by heme on enzymatic ALA synthesis, thus making ALA a very efficient PpIX prodrug widely used for clinical PDT, following either topical or systemic application (9). Nevertheless it was previously shown that exogenous application of PpIX to cells resulted in very efficient photocytotoxicity (10). One serious limitation in the exogenous use of PpIX much alike many other PSs is its low water solubility, and consequently the difficulty in preparing pharmaceutical formulations.…”

Influence of the photosensitizer photobleaching in the propagation of light during photodynamic therapy

“…Thus the initial photosensitizer absorption coefficient used as initial condition in the photochemical model (5)(6)(7)(8)(9)(10)(11) was obtained by means of equation 4, taking into account the previously obtained inhomogeneous photosensitizer concentration and an absorption cross-section of the PpIX, psa σ = 0.37·10 -15 cm 2 at 635 nm, which was derived from a study of the cellular photosensitizing properties of PpIX carried out in a transformed murine keratinocyte cell line 16 . Due to the fact that MAL is a derivative of 5-aminolevulinic acid, the relaxation time from singlet excited state to ground state S 1 to S 0 was set to 7.4 ns as reported earlier from fluorescence measurements in cells incubated with 5-aminolevulinic acid induced PpIX 17 .…”