Free and Protected Protease in the Diet of Lactating Jersey Cows: Effects on Performance, Milk Quality, Metabolism, Nutrient Digestibility, Microbiota, and Ruminal Environment
Maksuel Gatto de Vitt, Andrei Lucas Rebelatto Brunetto, Emeline Pizzolatto de Mello, Tainara Letícia dos Santos, Luisa Nora, Beatriz Danieli, Matheus Wroblescki Silva, Sander Souza Farias, Viviane Cargnin de Lima, Bruna Klein, Camila Ten Kathen Jung, Aniela Pinto Kempka, Gilberto Vilmar Kozloski, Roger Wagner, Miklos Maximiliano Bajay, Aleksandro Schafer da SilvaThis study evaluated the effects of dietary inclusion of free and protected acid protease on productive performance, milk composition, metabolic profile, nutrient digestibility, and ruminal environment in lactating Jersey cows. Fifteen multiparous cows (67 ± 7.5 days in milk; 27.5 ± 3.5 kg/day) were assigned to a 3 × 3 Latin square (5 squares) design with 21-day periods. Treatments consisted of: control (no enzyme), free protease (4.4 g/day), and protected protease (4.4 g/day). The protected form was developed using alginate-based encapsulation to enhance enzyme stability under ruminal conditions. Protease inclusion did not affect dry matter intake, milk yield, or feed efficiency (p > 0.05). However, free protease increased lactation persistency (p = 0.05) and improved fat-corrected and energy-corrected milk yields (p ≤ 0.02), with intermediate responses observed for protected protease. Milk fat and protein contents were higher in enzyme-fed cows (p ≤ 0.05), while other compositional parameters remained unchanged. Apparent crude protein digestibility was greater in cows receiving free protease (p = 0.037), with no effects on dry matter or fiber digestibility. Protease intake increased total volatile fatty acid concentrations and major fermentation products (acetate, propionate, and butyrate; p ≤ 0.01), indicating enhanced ruminal fermentation. Blood metabolites showed increased total protein and globulin levels in cows fed free protease (p ≤ 0.05), suggesting improved protein metabolism. Microbiota analysis revealed no differences in alpha or beta diversity; however, specific microbial taxa and predicted metabolic pathways were modulated by treatments, particularly in post-ruminal compartments. In conclusion, exogenous protease, especially in free form, improved protein utilization and corrected milk production without disrupting microbial stability. These findings highlight the potential of protease as a nutritional strategy to enhance efficiency in dairy systems through targeted modulation of ruminal function and nutrient metabolism.