Preparation of magnetic carbon nitride composite toward phosphopeptide enrichment

Protein phosphorylation plays an important role in cellular signaling and disease development. Advances in mass spectrometry-based proteomics have enabled qualitative and quantitative phosphorylation studies as well as in-depth biological explorations for biomarker discovery and signaling pathway analysis. However, the dynamic changes that occur during phosphorylation and the low abundance of target analytes render direct analysis difficult because mass spectral detection offers no selectivity, unlike immunoassays such as Western blot and enzyme-linked immunosorbent assay (ELISA). The present study aimed to solve one of the key problems in the specific and efficient isolation of phosphorylated peptides. A method based on a magnetic carbon nitride composite coupled with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was developed for the enrichment and analysis of phosphopeptides with low abundance in complex samples. Magnetic carbon nitride composite was synthesized and characterized by electron microscopy, infrared spectroscopy, and X-ray diffractometry. The composite showed a well-distributed two-dimensional layered structure and functional groups with excellent paramagnetic performance. Two classical phosphoproteins, namely,