Effect of negative dietary cation-anion difference (DCAD) diet on primiparous cow metabolomic analysis: Colostrum and milk assay.

Objectives: This study aimed to characterize the metabolomic profile of colostrum and milk in primiparous Holstein cows subjected to a prepartum diet with a low negative dietary cation-anion difference (DCAD). Materials and Methods: Eight primiparous cows were randomly assigned to two treatments: (a) basal diet (n = 4) and (b) basal diet supplemented with 250 gm/day of anionic salts (n = 4), starting 21 days before parturition. Colostrum and milk samples were analyzed using gas chromatography-mass spectrometry (GC-MS). Data were log-transformed and evaluated using multivariate (PLS-DA) and univariate (Wilcoxon test with fold change) approaches in MetaboAnalyst 6.0. Results: A total of 34 and 31 metabolites were detected in colostrum and milk, respectively. Multivariate analysis showed a visual separation between treatments; however, model validation parameters indicated limited predictive performance. Volcano plot analysis did not identify metabolites with statistical significance (p < 0.05), although several compounds exhibited consistent trends in relative abundance between groups. In colostrum, fatty acids such as arachidic acid and cholesterol tended to be lower in DCAD-supplemented cows, whereas L-glutamate and dodecanedioic acid showed higher relative abundance. In milk, similar trend-level differences were observed in fatty acid profiles between treatments. Conclusions: Prepartum supplementation with a negative DCAD diet was associated with exploratory metabolomic shifts in colostrum and milk of primiparous cows, particularly in lipid-related compounds. These findings should be interpreted as preliminary, and further studies with larger sample sizes are required to confirm the observed patterns.