Subclinical Hypocalcemia in Dairy Cows: An Integrative Evaluation of Blood Biomarkers, In-Line Milk Composition, and Rumination Behavior

Subclinical hypocalcemia (SCH) is one of the most prevalent metabolic disorders in early-lactation dairy cows, yet its multifaceted physiological effects are often overlooked due to the absence of clinical symptoms. This study aimed to characterize SCH through an integrative assessment of blood biochemical markers, in-line milk composition, and sensor-derived behavioral traits. Seventy-five Holstein cows between 2 and 21 days in milk were classified into hypocalcemic (group 1) (Ca < 2.0 mmol/L; <i>n</i> = 20) and healthy (group 2) groups (<i>n</i> = 55). Blood samples, milk data, and rumination metrics were evaluated, and group differences were analyzed using Welch's <i>t</i>-test and Pearson correlations. Cows with SCH exhibited significantly lower concentrations of Ca, PHOS, Mg, ALB, TP, GLUC, and Fe, indicating disruptions in mineral balance, protein metabolism, and energy status. Hepatic indicators (AST, ALT, GGT) did not differ between groups, whereas CREA was significantly lower in hypocalcemic cows, suggesting altered muscle metabolism rather than impaired renal function. Although differences in milk yield, composition, and rumination time did not reach statistical significance, hypocalcemic cows showed consistent biological tendencies toward reduced milk components and lower milk temperature. Correlation analysis revealed strong physiological linkages among Ca, ALB, P, TP, and Fe, underscoring the interconnected nature of mineral and protein metabolism in early lactation. These findings demonstrate that SCH is associated with coordinated biochemical and behavioral changes even in the absence of clinical signs. Integrating blood biomarkers with real-time sensor data provides a more comprehensive understanding of calcium-related metabolic challenges and highlights the potential of precision-livestock technologies for early detection. Future studies incorporating ionized calcium and longitudinal sampling are needed to refine diagnostic thresholds and improve predictive monitoring of SCH in dairy herds.