AbstractA novel hydrazide functionalized magnetic chitosan nanostructure was developed as a sorbent for the selective extraction of peptides with N‐terminal serine and threonine. Iron oxide nanoparticles were synthesized by the co‐precipitation method. Chitosan was covalently bonded onto the surface of magnetic Fe3O4 nanoparticles. Magnetic chitosan was modified by glutaraldehyde as a linking agent. Hydrazide functional groups were created on the sorbent surface by immobilization of adipic dihydrazide. The sorbent characterization was performed by scanning electron microscopy and Fourier transform infrared spectroscopy. The studied peptides were labeled by oxidation of the hydroxyl group in 1,2‐amino alcohol structure under controlled conditions. The aldehyde‐form peptides were immobilized on the surface of the sorbent through the hydrazide group. The formed hydrazone compounds were eluted by hydroxylamine reagent. The release oximated peptides were determined by high‐performance liquid chromatography. The effect of extraction time, elution time and elution volume on extraction performance were studied. Extraction and desorption processes were performed in 6 h. The adsorption efficiency of the sorbent was 68 and 61% for peptides with N‐terminal serine and threonine, respectively. The sorbent showed excellent specificity in extracting peptides with N‐terminal serine and threonine in the presence of a peptide with non‐N‐terminal serine and threonine.