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Engineered nanomaterials (ENMs) are key drivers for the development of highly sophisticated new technologies. As all new attainments, the rapidly increasing used of ENMs raise concerns about their safety for the environment and humans. There is growing evidence showing that if engineered nanomaterials are released into the environment, there is a possibility that they could cause harm to aquatic microorganisms. Among the divers effects triggering their toxicity the ability of ENMs to generate reactive oxygen species (ROS) capable of oxidizing biomolecules is currently considered a central mechanism of toxicity. Therefore, development of sensitive tools for quantification of the ROS generation and oxidative stress are highly sought. After briefly introducing ENMs-induced ROS generation and oxidative stress in the aquatic microorganisms (AMOs), this overview paper focuses on a new optical biosensor allowing sensitive and dynamic measurements of H 2 O 2  in real-time using multiscattering enhanced absorption spectroscopy. Its principle is based on sensitive absorption measurements of the heme protein cytochrome  c  whose absorption spectrum alters with the oxidation state of constituent ferrous Fe II  and ferric Fe III . For biological applications cytochrome  c  was embedded in porous random media resulting in an extended optical path length through multiple scattering of light, which lowers the limit of detection to a few nM of H 2 O 2 . The sensor was also integrated in a microfluidic system containing micro-valves and sieves enabling more complex experimental conditions. To demonstrate its performance, abiotic absorption measurements of low concentrations of dye molecules and 10 nm gold particles were carried out achieving limits of detection in the low nM range. Other biologically relevant reactive oxygen species can be measured at sub-μM concentrations, which was shown for glucose and lactate through enzymatic reactions producing H 2 O 2 . In ecotoxicological investigations H 2 O 2  excreted by aquatic microorganisms exposed to various stressors were measured. Pro-oxidant effects of nano-TiO 2  and nano-CuO towards green alga  Chlamydomonas reinhardtii  were explored in various exposure media and under different light illuminations. Dynamics of Cd 2+  induced effects on photosynthetic activity, sensitisation and recovery of cells of  C. reinhardtii  was also studied.

Non-invasive continuous monitoring of pro-oxidant effects of engineered nanoparticles on aquatic microorganisms