This paper was presented at the 4th Micro and Nano Flows Conference (MNF2014), which was held at University College, London, UK. The conference was organised by Brunel University and supported by the Italian Union of Thermofluiddynamics, IPEM, the Process Intensification Network, the Institution of Mechanical Engineers, the Heat Transfer Society, HEXAG - the Heat Exchange Action Group, and the Energy Institute, ASME Press, LCN London Centre for Nanotechnology, UCL University College London, UCL Engineering, the International NanoScience Community, www.nanopaprika.eu.The study of reactive oxygen species (ROS) generation caused by nano-bubbles (NBs) is of great importance for its application in both physiological activity promotion aspect and sterilization aspect. In this paper, Aminophenyl Fluorescein (APF) was used as a fluorescent reagent for the detection of ROS generation by NBs. The NBs scattering could cause the decrease of fluorescence intensity. Meanwhile, the quenching effect of oxygen could also cause the decrease of fluorescence intensity. Although the above two factors masked the fluorescence intensity generation by ROS, the fluorescence intensity of the water containing NBs still increased with NBs generation time, which proved that oxygen NBs could generate ROS. Moreover, the endogenous ROS in the barley seed cells were measured in the seed that germinated in the water containing NBs and the distilled water respectively. According to the results of seed staining experiments, both the microscope images and the absorbance at 560nm proved that the seeds dipped in the water containing NBs could generate more ROS compared to those in the distilled water. These findings greatly increase the NBs potential use both in agricultural field and environmental field