Preparation process optimization of pig bone collagen peptide-calcium chelate using response surface methodology and its structural characterization and stability analysis

Abstract

In this study, alcalase and neutrase were used in combination to prepare collagen peptides with high calcium binding ability. The optimal conditions for the preparation of peptide-calcium chelate (mass ratio of peptide/calcium of 4.5:1 for 40 min at 50 °C and pH 9) were determined by response surface methodology (RSM), under which a calcium chelating rate of 78.38% was obtained. The results of Ultraviolet-Visible (UV-Vis), fluorescence and Fourier transform infrared (FT-IR) spectra synthetically indicated that calcium could be chelated by carboxyl oxygen and amino nitrogen atoms of collagen peptides, thus forming peptide-calcium chelate. The chelate was stable at various temperatures and pH values, and exhibited excellent stability in the gastrointestinal environment, which could promote calcium absorption in human gastrointestinal tract. Moreover, Caco-2 cell monolayer model was used to investigate the effect of peptide-calcium chelate on promoting calcium absorption. Results showed that peptide-calcium chelate could significantly improve calcium transport in Caco-2 cell monolayer and reverse the inhibition of calcium absorption by phosphate and phytate. The findings provide a scientific basis for developing new calcium supplements and the high-value utilization of pig bone.

Keywords: Absorption; Caco-2 cell monolayer model; Calcium dichloride (PubChem CID: 5284359); Calcium-binding peptide; Cholecalcifetrol (PubChem CID: 5280795); Citric acid (PubChem CID: 311); Ethanol (PubChem CID: 702); Formaldehyde (PubChem CID: 712); Hexane (PubChem CID: 8058); Peptide-calcium chelate; Pig bone; Potassium Phosphate, Monobasic (PubChem CID: 516951); Response surface methodology; Sodium hydroxide (PubChem CID: 14798); Sodium phytate (PubChem CID: 66391); Structural characterization; Trichloroacetic acid (PubChem CID: 6421).