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Chitosan (CTS)/chitosan oligosaccharide (COS) and whey protein isolate (WPI) have been frequently used as food supplements, but notably, the interaction between the carbohydrate and the protein may affect the digestibility of protein. Thus, the present study focused on effects of the interaction between CTS/COS and WPI on the protein digestibility. A series of chemical and spectroscopic techniques including gel electrophoresis, gel permeation chromatography, Fourier transform-infrared (FT-IR) spectroscopy, intrinsic fluorescence (IF) spectroscopy, and circular dichroism (CD) spectroscopy were applied. According to the findings, both CTS and COS dramatically reduced intestinal digestibility of WPI, resulting in a decrease of DH by 43.33 % and 52.31 %, respectively. The substitution degree of WPI on CTS was 0.87 g WPI/g CTS, and the electrostatic interaction between amine groups of CTS and carboxyl groups of WPI caused changes in WPI's stability, microstructure, and fluorescence intensity. Notably, CTS affected the digestibility of WPI by precipitating protein and enzyme, whereas COS altered WPI's digestibility by decreasing or inactivating enzyme activity. The present study offered a solid scientific foundation for the rational formulations of carbohydrates and proteins in food industry. [Display omitted] • Chitosan (CTS) and its oligosaccharide (COS) influenced protein digestibility. • CTS decreased protein digestibility by partially precipitating protein and protease. • COS influenced protein digestibility by decreasing the activity of protease. • The electrostatic interaction occurred between -NH 3 + of CTS and -COO− of protein. [ABSTRACT FROM AUTHOR]