Colostrum is particularly rich in proteins and bioactive peptides that play an important role in the health of newborns. Proteomics can help identify and characterise these components, which is crucial for understanding their effects on newborns. The study material consisted of colostrum collected 12 h after delivery from six primiparous Polish noble half-bred mares. Two-dimensional electrophoresis and mass spectrometry were used to separate, identify and quantify mare colostrum whey proteins and peptides. The analysis revealed that the largest group comprised extracellular proteins (49% of the total), followed by cytoplasmic proteins (31%). The biological processes in which the identified proteins were primarily involved included cellular component organisation, response to stress, establishment of localisation, defence response, plasma membrane-bound cell projection organisation, cell projection organisation, regulation of protein metabolic process, response to biotic stimulus. The proteins were primarily associated with the following molecular functions: small molecule binding, ion binding, molecular function regulator activity, cation binding, enzyme regulator activity, carbo-hydrate derivative binding, enzyme binding, signalling receptor binding, calcium ion binding, lipid binding, cytoskeletal protein binding, molecular function activator activity. Knowledge of the protein composition of mare colostrum whey is crucial for understanding the role of colostrum in the healthy development of newborns. Due to differences in colostrum protein composition between individual horse breeds, further proteomic research is needed to characterise these differences and assess their implications.