Phosphatidylcholine oxidation is closely related to many neurodegenerative diseases. In this paper, Raman spectroscopy was proposed to continuously monitor the oxidation of phosphatidylcholine and provide deep understanding of this biochemical process. To increase the detection sensitivity, surface-enhanced Raman spectroscopy (SERS) with a micro-nanosilver-complex substrate was prepared by electrodeposition. The prepared SERS substrate had an enhancement factor as high as 7.8 × 107, ensuring detection sensitivity in the phosphatidylcholine-oxidation process. It was illustrated that the oxidation of phosphatidylcholine in an ethanol-water solution under the experimental conditions could be monitored and well described by second-order kinetics by continuously measuring and analyzing the SERS spectra of phosphatidylcholine-oxidation intermediates in 20 days. Meanwhile, the oxidation products were confirmed by mass spectrometry, and the oxidation process was in good concordance with mass-spectrometry detection. The use of SERS in following a biochemical process has advantages, including simple instrumentation, a low cost, a short detection time, and no sample pretreatment. Therefore, as a kind of vibration spectrum, SERS is preferable to traditional detection approaches such as MS, HPLC, and MRI for the dynamic monitoring and analysis of complex biochemical processes.