The ROS seemed to be endlessly produced by P25TiO₂NPs upon irradiation, since the values detected after 6 h are similar to the ones after 3 h. However, the amount of vitamin B2 in the surface of the NPs proved to be enough to decrease the ROS detected even after 6 h. Statistical analysis showed a significant difference between C and A. p < 0.05

The skin of an adult person is, in most places, covered with a relatively thick (∼10 μm) barrier of keratinised dead cells. One of the main questions is still whether TiO₂ NPs are able to penetrate into the deeper layers of the skin. The majority of studies suggest that TiO₂ NPs, neither uncoated nor coated (SiO₂, Al₂O₃ and SiO₂/Al₂O₃) of different crystalline structures, penetrate normal animal or human skin. However, in most of these studies the exposures were short term (up to 48 h); only few long-term or repeated exposure studies have been published. Wu et al.83 have shown that dermal application of nano-TiO₂ of different crystal structures and sizes (4–90 nm) to pig ears for 30 days did not result in penetration of NPs beyond deep epidermis. On the other hand, in the same study the authors reported dermal penetration of TiO₂ NPs with subsequent appearance of lesions in multiple organs in hairless mice, that were dermal exposed to nano-TiO₂ for 60 days. However, the relevance of this study for human exposure is not conclusive because hairless mice skin has abnormal hair follicles, and mice stratum corneum has higher lipid content than human stratum corneum, which may contribute to different penetration. Recently Sadrieh et al. performed a 4 week dermal exposure to three different TiO₂ particles (uncoated submicron-sized, uncoated nano-sized and coated nano-sized) in 5 % sunscreen formulation with minipigs. They found elevated titanium levels in epidermis, dermis and in inguinal lymph nodes, but not in precapsular and submandibular lymph nodes and in liver. With the energy dispersive X-ray spectrometry and transmission electron microscopy (TEM) analysis the authors confirmed presence of few TiO₂ particles in dermis and calculated that uncoated nano-sized TiO₂ particles observed in dermis represented only 0.00008 % of the total applied amount of TiO₂ particles. Based on the same assumptions used by the authors in their calculations it can be calculated that the total number of particles applied was 1.8 × 10¹³ /cm² and of these 1.4 x10⁷/cm² penetrated. The surface area of skin in humans is around 1.8 m²  and for sun protection the cream is applied over whole body, which would mean that 4 week usage of such cream with 5 % TiO₂ would result in penetration of totally 2.6 × 10¹⁰ particles. Although Sadrieh et al.concluded that there was no significant penetration of TiO₂ NPs through intact normal epidermis, the results are not completely confirmative.