The objective of this study was to compare the properties of small intestinal fluid (SIF) from white- and yellow-feathered broilers fed variable dietary CP to test the hypothesis that digestive enzymes sourced from the SIF of yellow-feathered broilers could serve as a viable alternative for preparing in vitro SIF specifically for white-feathered broilers. Ten Arbor Acres Plus broilers (32 days of age) and 20 Chinese yellow chickens (51 days of age) were fitted with jejunal cannulas and assigned to dietary treatments as follows: 1) white-feathered broilers (BW of 4.08 ± 0.39 kg) fed diet 1 with CP of 19.88 %, 2) yellow-feathered broilers (BW of 2.89 ± 0.27 kg) fed diet 1, and 3) yellow-feathered broilers (BW of 2.84 ± 0.20 kg) fed diet 2 with CP of 16.32 %. Each treatment contained 10 replicates with 1 broiler in each replicate. Digestive enzyme activities, ion concentrations, pH, 16 amino acid (AA, excluding Tyr and Trp) contents, molecular weight distribution (MWD) of protein, hydrolysis rates on wheat starch, corn starch, casein, and soybean protein concentrate, and in vitro digestible energy of corn, soybean meal, and corn gluten meal were evaluated for SIF. Activities of chymotrypsin and amylase, pH, concentrations (mg/mL) of total amino acid (TAA) and 13 AA (excluding Pro, Cys, and Glu), and hydrolysis rates in casein or soybean protein concentrate were greater (P < 0.05), whereas MWD of protein from 19 to 23 kDa was lower (P < 0.05) in the SIF of yellow compared to white-feathered broilers fed diet 1. Furthermore, activities of chymotrypsin and amylase, concentrations of Na+, and Cl−, MWD of protein from 15 to 18 kDa were greater, while concentrations of K+ were lower in the SIF of yellow-feathered broilers fed diet 2 compared to white feathered broilers fed diet 1. Activities of chymotrypsin and amylase, K+ concentration, pH, TAA and 13 AA (excluding Pro, Cys, and Glu), hydrolysis rates in casein or soybean protein concentrate increased (P < 0.05), while concentrations of Na+ and Cl− (P < 0.05) decreased with dietary CP in SIF from yellow-feathered broilers. This experiment demonstrated diet composition and breed modulate digestive enzyme activities, ions (Na+, K+, Cl−), pH, and AA concentrations in the SIF. Our findings revealed no substantial difference in the hydrolytic characteristics of SIF from white-feathered broilers fed diet 1 and yellow-feathered broilers fed diet 2. Consequently, it is feasible to use digestive enzymes extracted from the SIF of yellow-feathered broilers to prepare simulated SIF for in vitro digestion of white-feathered broilers.
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