Low-dose-rate ionizing radiation increases singlet oxygen production by photosensitizers

Photosensitizers have significant potential as radiosensitizers in cancer treatment, yet the mechanism of ionizing-radiation-induced singlet oxygen (¹O₂) generation remains unclear. Here, we systematically investigated ¹O₂ production by the photosensitizer Chlorin e6 (Ce6) using the Singlet Oxygen Sensor Green probe and imidazole/ p -nitroso- N , N -dimethylaniline detection methods, evaluating the effects of photon energy (X-rays up to 310 kV and ⁶⁰Co gamma rays at 1.17 and 1.33 MeV), dose, and dose rate. Ce6 produced more ¹O₂ with increasing photon energy. At 5 Gy, the lowest dose rate (0.005 Gy/min) yielded significantly more ¹O₂ than higher dose rates (7–0.05 Gy/min). Scavenging experiments identified superoxide anions (·O₂⁻) as a key intermediate. We propose that, unlike classical triplet-state photosensitization, ionizing radiation induces Ce6 radical cations (Ce6^⋅+), which react with radiation-induced ·O₂⁻ to generate ¹O₂. These findings suggest potential for photosensitizer-radiation combinations in low-dose-rate therapies, although further biological validation and consideration of tumor redox status are required.