The toxicity of P25TiO₂NPs was evaluated in both prokaryotic (Fig. 3) and eukaryotic cells (Fig. 4). The XTT assay was chosen to measure the cell viability in bacterial cultures of MSSA, a normal skin microbiota microorganism. The reduction in the viability of samples with bare NPs is notorious, possibly due to the described ROS production from the interaction of P25TiO₂NPs with light [37]. This effect seems to be avoided when they are functionalized with vitamin B2. Also, the most concentrated vitaminB2@P25TiO₂NPs sample (0.2 mg/mL) shows up to 60% more absorbance after 6 h compared to the bare NPs (due to normal cell replication). This may indicate that the antioxidant effect of the vitamin B2 coating is greater than the oxidation damage produced by the NPs. This protective capacity could be attributed to the glutathione redox cycle and the conversion of reduced riboflavin to its oxidized form [38]. Values of cell viability greater than 100% are not rare and could be understood because the XTT assay actually measure metabolic activity when reducing the tetrazole to formazan. It is usually assumed that conversion is dependent on the number of viable cells, but it could also be related to an expected increased enzymatic activity when cells are exposed to small doses of some new substance. Further analysis showed that this effect was not the only one responsible for better cell viability of vitaminB@P25TiO₂NPs treated samples.

What is titanium dioxide? Why is it used in food products?

Method of producing improved lithopone

Nano-sized TiO₂ generally shows low or no acute toxicity in both invertebrates and vertebrates. However, exposure of Daphnia magna to 20 ppm TiO₂ for 8 consecutive days was found to cause 40 % mortality. Zhu et al. showed minimal toxicity to D. magna after 48 h exposure, while upon chronic exposure for 21 days, D. magna suffered severe growth retardation and mortality. A significant amount of nano-sized TiO₂ was found also accumulated in the body of the animals. Similar findings with coated nano-sized TiO₂ (T-Lite™ SF, T-Lite™ SF-S and T-Lite™ MAX; BASF SE) were reported by Wiench et al. Biochemical measurements showed that exposure to TiO₂ NPs induces significant concentration-dependent antioxidant enzyme activities in D. magna. Lee et al. showed that 7 and 20 nm-sized TiO₂ induced no genotoxic effect in D. magna and in the larva of the aquatic midge Chironomus riparius.

Australian researchers examined how titanium dioxide as a food additive affected gut microbiota in mice by orally administering it in drinking water. The study, published in the journal Frontiers in Nutrition in 2019, found the treatment could “alter the release of bacterial metabolites in vivo and affect the spatial distribution of commensal bacteria in vitro by promoting biofilm formation. We also found reduced expression of the colonic mucin 2 gene, a key component of the intestinal mucus layer, and increased expression of the beta defensin gene, indicating that titanium dioxide significantly impacts gut homeostasis.” The changes were then linked to colonic inflammation, along with a higher expression of inflammatory cytokines, which are signal proteins that help with regulation. The researchers concluded that titanium dioxide “impairs gut homeostasis which may in turn prime the host for disease development.”

Health Canada's Food Directorate recently completed a “state of the science” report on titanium dioxide (TiO₂) as a food additive. Food-grade TiO₂ is a white powder made up of small particles that has been permitted in Canada and internationally for many years as a food additive to whiten or brighten foods. Food-grade TiO₂ has long been considered safe in Canada and in other countries when eaten as part of the diet.

TiO₂ comes in many different forms. However, only a few of these forms are considered food-grade (acceptable to be added to food). Many studies that raised concern about the safety of TiO₂, including the concern for genotoxicity, used forms of TiO₂ that are not considered acceptable for use in food and have different properties than food-grade TiO₂. Other studies did use food-grade TiO₂, but took steps to break the material down into smaller particles than what would normally be found in food.