Effect of TiO2 particle size: haemolytic activity and cell viability determination interference

Abstract

It is well known that ultraviolet radiation harms the skin and can lead to skin cancer. The correct use of sunscreen is the most efficient way to prevent the development of this disease. The photoprotection characteristic of sunscreens is mainly attributed to their constituent UV filters, which can be organic or inorganic. A frequently used inorganic filter is TiO2, both in its micro and nano forms. Micro-TiO2 has a whitening effect in sunscreen formulations, decreasing their aesthetic appeal, which can be resolved by using the nano form of TiO2. Due to the greater surface area/volume ratio, nanoparticles become more (bio)reactive giving rise to concerns about their potential toxicity, for instance generating Reactive Oxygen Species (ROS) known to be implicated in cellular damage. This study compares the toxic behaviour of two commercially available TiO2 at a micro and nanoscale. For this purpose, the toxicity of this compound on HaCaT epidermal keratinocytes is assessed. Prior to these cytotoxicity studies, the potential interferences when using colorimetric methods (MTT, NRU and LDH) needed to be discarded. Additionally, the haemolytic effect of TiO2 on human red blood cells is studied and nano-TiO2 is characterized through TEM. Results show that neither micro-TiO2 nor nano-TiO2 exhibit considerable haemolysis on red blood cells, even though nanoparticles display higher haemolysis values. The particle size of nano-TiO2 has been confirmed by TEM. No chemical interferences of TiO2 with the reactants of the MTT, NRU or LDH methods are found, although the highly insoluble TiO2 particles could interfere with the absorbance measurement. Finally, TiO2 does not exhibit cytotoxicity to HaCaT epidermal keratinocytes for NRU and LDH, although a reduction in cell viability in MTT for NPs was observed.