Mass Spectrometric Analysis of the Photobleaching of Protoporphyrin IX Used in Photodynamic Diagnosis and Therapy of Cancer

Abstract

Photobleaching and photoproduct formations are considered essential phenomena in improving the efficacy of photodynamic diagnosis and therapy (PDD and PDT). We investigated the photobleaching of protoporphyrin IX (PpIX) by measuring its concentration with mass spectrometry (MS). The reduction in the concentration of PpIX dissolved in dimethyl sulfoxide was measured during PDD and PDT conditions using lasers with wavelengths of 405 and 635 nm, respectively, at a power density of 10, 50 or 100 mW/cm2. The obtained results were compared with the results of conventional fluorescence spectroscopy and previously reported results. Our results demonstrate the variation in the MS‐based photobleaching coefficient of PpIX with the power density, while the fluorescence‐based photobleaching coefficient was independent of the power density. The results of MS also show faster photobleaching of PpIX in comparison with that obtained from fluorescence. The difference may be attributed to the change in the fluorescence quantum yield of PpIX with its concentration and the effect of fluorescence emission from the PpIX photoproducts. Thus, an MS‐based investigation of the photobleaching poses to be a more stable investigation form. Our finding highlights the importance of recognizing the potential significance of these discoveries in the PDD and PDT dosimetry and efficacy.

Increased fluorescence observation intensity during the photodynamic diagnosis of deeply located tumors by fluorescence photoswitching of protoporphyrin IX

“…The spectroscopic analysis indicated that the rate at which PpIX photodegraded at the 633 nm peak did not vary with power density for either excitation wavelengths. This result is in line with our previous study of PpIX photobleaching under 405 and 635 nm irradiation 14 . The consistent photobleaching rate of ALA-induced PpIX over a range of power densities in tumor models and mouse skin in vivo has also been reported 31 , 32 .…”

“…This result is in line with our previous study of PpIX photobleaching under 405 and 635 nm irradiation. 14 The consistent photobleaching rate of ALA-induced PpIX over a range of power densities in tumor models and mouse skin in vivo has also been reported. 31,32 On the other hand, a discrepancy was seen in the findings reported by others, in which PpIX photobleaching was dependent on the power density.…”

“…The concentration and, thus, the fluorescence intensity of the photosensitizer reduce under light exposure, decreasing the observed fluorescence. 4,14,15 Aside from the wavelength of light used in the irradiation, the photobleaching rate of PpIX is dependent on its initial concentration. [16][17][18] Goyan et al observed a 50% reduction of 10 μM PpIX solution after 25 min of 300 mW irradiation with 764 nm light.…”

“…During the fluorescence diagnosis process in PDD, the photobleaching of the photosensitizer occurs. The concentration and, thus, the fluorescence intensity of the photosensitizer reduce under light exposure, decreasing the observed fluorescence 4 , 14 , 15 . Aside from the wavelength of light used in the irradiation, the photobleaching rate of PpIX is dependent on its initial concentration 16 …”

Recent Advances in the HPPH-Based Third-Generation Photodynamic Agents in Biomedical Applications

“…During photodynamic reactions, certain photosensitizers can also react with generated singlet oxygen ( 1 O 2 ), resulting in decreased photosensitizer concentration, absorbance, and fluorescence intensity. This phenomenon is termed photobleaching [56]. Due to the spectral characteristics of HPPH absorption, exposure to visible light after injection or fluorescence imaging prior to treatment may prompt the photosensitizer (PS) to reach an excited state prematurely, before its intended therapeutic activation.…”