Smart Ratiometric SERS Nanoprobe for Real-Time Monitoring Hydrogen Peroxide in Living Cells during NADH Treatment Associated with Ferroptosis

Studying the oxidative stress, especially the reactive oxygen species (ROS) response of ferroptosis, is crucial for the diagnosis and treatment of cancer based on ferroptosis. However, reliable quantitative analysis of intracellular ROS in cancer treatment for drug screening is still a challenge. Herein, a superior ratiometric SERS nanoprobe was developed for in situ, real-time, and highly sensitive detection of content variation of H₂O₂ within living cells. The SERS nanoprobe was prepared by coassembly of the internal standard molecule p-mercaptobenzonitrile and the reporter molecule p-mercaptophenylboronic acid on the surface of gold nanoparticles, used for synergistic calibration and detection of H₂O₂, which enables reliable detection of the true content of intracellular H₂O₂ without the interference of other substances in cells. Based on the nanoprobe, we found that the level of intracellular H₂O₂ of cancer cells was increased after the nicotinamide adenine dinucleotide (NADH) treatment, with a dose-dependence to the concentration of NADH. High doses of NADH (above 20 mM) can induce cell death by means of ferroptosis associated with the level elevation of intracellular lipid hydroperoxides. This study highlights the potential of the SERS nanoprobe for tracking content variation of cellular H₂O₂ and understanding its roles in screening new anticancer drugs.