A novel static affinity chromatography strategy for efficient isolation of cod bone calcium-binding peptide and evaluation of its chelation mechanism and calcium transport.

“A novel static affinity chromatography strategy for efficient isolation of cod bone calcium-binding peptide and evaluation of its chelation mechanism and calcium transport. Tian Q(1), Hao L(2), Fan C(2), Hou H(3). Author information: (1)State Key Laboratory of Marine Food Processing & Safety Control, College of Food Science and Engineering, Ocean University of China, No.1299, Sansha Road, Qingdao, Shandong Province 266404, PR China; Yantai Institute of Coastal Zone Research, Chinese Academy of Sciences, Yantai, Shandong Province 264003, PR China. (2)State Key Laboratory of Marine Food Processing & Safety Control, College of Food Science and Engineering, Ocean University of China, No.1299, Sansha Road, Qingdao, Shandong Province 266404, PR China. (3)State Key Laboratory of Marine Food Processing & Safety Control, College of Food Science and Engineering, Ocean University of China, No.1299, Sansha Road, Qingdao, Shandong Province 266404, PR China; Laboratory for Marine Drugs and Bioproducts, Qingdao Marine Science and Technology Center, No.168, Wenhai Middle Road, Jimo District, Qingdao, Shandong Province 266237, PR China; Sanya Oceanographic Institution, Ocean University of China, Sanya, Hainan Province 572024, PR China. Electronic address: houhu@ouc.edu.cn. The low yield of high-activity calcium-binding peptides constitutes a technical bottleneck restricting their application. Herein, a static affinity chromatography approach using needle-shaped hydroxyapatite was developed for the isolation of a novel cod bone calcium-binding peptide, GRGNEGPQ (CBP-8). The calcium-binding mechanism was investigated using experimental and computational methods, while its digestive behavior and calcium transport effects were evaluated using an in vitro digestion model and Caco-2 cell monolayer model. Results indicated that the carboxyl group of Gln served as a potential calcium-binding site and the CBP-8-Ca complex formed through multiple chelation modes. Electrostatic interactions were critical for coordination bond formation, with a formal charge transfer of -2.13 e from Ca2+ in the CBP-8-Ca complex. CBP-8-Ca exhibited good gastric stability but degraded intestinally. Additionally, CBP-8 promoted Ca2+ transport via non-specific calcium channels and TRPV6 channels. This study offers a promising and cost-effective strategy for the efficient production of highly active calcium-binding peptides. Copyright © 2026 Elsevier Ltd. All rights reserved. Conflict of interest statement: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.”