The fluorescence measurements in Figure 1b,c shows that all the specific ROS increased with the irradiation time, but the N-TiO2 induced more O2  ·−/H2O2 (Figure 1b) while less OH · (Figure 1c) than TiO2. It was reported that the photogenerated holes of N-TiO2 were trapped in the N 2p levels and had a very low mobility [26], thus were barely involved in the photocatalysis when the N-TiO2 was illuminated by visible light [27]. In this study, the lower production of OH · from N-TiO2 might result from the same reason. However, the photogenerated

electrons in the conduction band can react with oxygen molecules to generate O2  ·−, which is thermodynamically favored [28]. Thus, N-TiO2 could generate more O2  ·−/H2O2 than the pure TiO2 CHIR98014 datasheet due to the AZD2014 higher visible light absorption efficiency. When cells were treated with TiO2 or N-TiO2 nanoparticles, the nanoparticles were not only found on the cell membrane but also in the cytoplasm, and some of them aggregated around or in Golgi complexes and even in nuclei [10]. As the TiO2 or N-TiO2 nanoparticles can induce ROS under visible light irradiation, the photokilling effect on cancer cells was observed in our previous work [10]. Considering that the productions of the specific ROS species generated by TiO2 or N-TiO2 are different and the contributions from the specific ROS to PDT may also be different, the PDT-induced changes of the intracellular

parameters, such as MMP, Ca2+, and NO concentrations in HeLa cells treated with TiO2 or N-TiO2 were studied as follows. MMP changes When TiO2- or N-TiO2-treated cells were illuminated by light, the generated ROS may attack the mitochondria [29] or the activated nanoparticles may interact ARRY-438162 with the mitochondria directly [30], which would affect the

function of mitochondria and cause the opening of mitochondrial permeability pores, resulting in the dissipation of MMP [30–32]. In this study, the MMP decreased immediately after the PDT as shown in Figure 2. It seems that the mitochondrion is a very sensitive cellular organelle during the PDT, and the defects can be detected immediately in our study. For TiO2-treated cells, the MMP level decreased continuously after the PDT with an approximate rate of 1.2% per min within 60 min. The MMP level for N-TiO2 samples dropped much faster (around 4.2% per min) O-methylated flavonoid within the first 10 min after the PDT, then decreased at slower and slower rate within 45 min, and almost kept in a constant rate of 20% after 45 min. However, the MMP levels of control cells and the cells incubated with TiO2 and N-TiO2 under light-free conditions did not show any change during 60 min (data not shown), which confirmed the low cytotoxicity of TiO2 and N-TiO2. Figure 2 MMP of HeLa cells as a function of the time after the PDT. Cells were incubated with 100 μg/ml TiO2 (white square) or N-TiO2 (black circle) for 2 h and illuminated by the visible light for 5 min. The averaged fluorescence intensity of control cells (white triangle) at 0 min was set as 100%.