Donkey milk has been widely shown to be an ideal substitute for human milk because of its similar composition. However, alterations to the composition of donkey milk during lactation have not been well studied. In this study, untargeted metabolomics with ultra-high-performance liquid tandem chromatography quadrupole time-of-flight mass spectrometry were used to analyze and compare the metabolites in donkey colostrum (DC) and mature milk (DMM). Two hundred seventy metabolites were characterized in both DC and DMM. Fifty-two of the metabolites in the DC were significantly different from those in the DMM; 8 were downregulated and 44 were upregulated. This demonstrated that the composition of the donkey milk changed with lactation. Additionally, the interactions and metabolic pathways were further analyzed to explore the mechanisms that altered the milk during lactation. Our results provide comprehensive insights into the alterations in donkey milk during lactation. The results will aid in future investigations into the nutrition of donkey milk and provide practical information for the dairy industry.

Cow milk (CM) allergy is one of the most common food allergies worldwide; the most abundant CM proteins, such as casein (CN), β‐lactoglobulin (β‐LG), and ɑ‐lactalbumin (ɑ‐LA), are all potentially allergenic. Reducing the antigenicity of CM continues to be a major challenge. However, previous studies have focused on the antigenicity of individual allergic CM proteins. Thus, in the present study, we aimed to evaluate the effects of different food‐grade enzymes on the antigenicity of CN, β‐LG, ɑ‐LA in natural CM. The degree of hydrolysis (DH) and molecular mass (MW) distribution of CM hydrolysates were assessed. Additionally, the residual antigenicity of CM hydrolysates was evaluated through enzyme‐linked immunosorbent assay and Western blotting with anti‐CN, anti‐β‐LG, and anti‐ɑ‐LA rabbit polyclonal antibodies. The results showed that Alcalase‐ and Protamex‐mediated hydrolysis could efficiently reduce the antigenicity of CN, β‐LG, and ɑ‐LA, inducing a higher DH, the loss of density of CM proteins, and the increasing levels of low MW (<3 kDa) peptides in CM hydrolysates. Further, Protamex and Alcalase could more efficiently hydrolyze the major allergenic components of CM than the other enzymes, which could represent an advantage for the development of hypoallergenic CM. These findings add further knowledge about the study and development of hypoallergenic CM.