TitleMass spectral profiling of caseinomacropeptide extracted from feeding material and jejunal fluid using three methods-ethanol precipitation, perchloric acid precipitation, and ultrafiltration.
Publication TypeJournal Article
Year of Publication2023
AuthorsKoh, J, Kim, BJin, Qu, Y, Dallas, DC
JournalFood Chem
Date Published08/2022
KeywordsAnimals, Caseins, Cattle, Cytidine Monophosphate, Ethanol, Humans, Jejunum, Peptide Fragments, Perchlorates, Ultrafiltration


  • Profiles of caseinomacropeptide (CMP) varied depending on the extraction method.
  • Ethanol precipitation was optimal for intact CMP extraction in feed and jejunal fluid.
  • Perchloric acid was optimal for fragment CMP extraction in jejunal fluid.
  • Ultrafiltration resulted in significant losses of peptides from the feeding material.


The ability of bovine κ-casein-derived caseinomacropeptide (CMP) to exert bioactivity in the human gut depends on its digestive survival. Sampling from the human jejunum after feeding CMP and top-down glycopeptidomics analysis facilitates the determination of CMP survival. To reduce interference from non-target molecules in mass spectrometric analysis, CMP must be isolated from digestive fluid. To identify an optimal extraction method, this study compared the profiles of CMP extracted from feeding material (commercial CMP in water) and digestive fluid by ethanol precipitation, perchloric acid (PCA) precipitation, and ultrafiltration. Ethanol precipitation yielded the highest ion abundances for aglycosylated CMP and glycosylated CMP in both feeding material and jejunal samples. Notably, PCA precipitation yielded the highest abundance of partially digested CMP-derived fragments in jejunal samples. Overall, ethanol precipitation was the most effective among the methods tested for intact CMP extraction from jejunal fluids, whereas PCA precipitation was optimal for extraction of CMP fragments.

Alternate JournalFood Chem
PubMed ID35969996