Identification of bitter peptides in aged Cheddar cheese by crossflow filtration-based Fractionation, Peptidomics, statistical screening and sensory analysis.

Despite bitterness being a common flavor attribute of aged cheese linked to casein-derived peptides, excessive bitterness is a sensory flaw that can lead to consumer rejection and economic loss for creameries. Our research employs a unique approach to identify bitter peptides in cheese samples using crossflow filtration-based fractionation, mass spectrometry-based peptidomics, statistics and sensory analysis. Applying peptidomics and statistical screening tools, rather than traditional chemical separation techniques, to identify bitter peptides allows for screening the whole peptide profile. Five peptides-YPFPGP (ß-casein [60-65]), YPFPGPIPN (ßA2-casein [60-68]), LSQSKVLPVPQKAVPYPQRDMPIQA (ß-casein [165-189]), YPFPGPIHNS (ßA1-casein [60-69]) and its serine phosphorylated version YPFPGPIHN[S] (ßA1-casein [60-69])- demonstrated high levels of bitterness with mean bitterness intensity values above 7 on a 15-point scale. In the future, this data can be combined with the microbial and protease profile of the Cheddar samples to help understand how these factors contribute to bitter taste development.

A comprehensive database of cheese-derived bitter peptides and correlation to their physical properties.

Bitterness is a common flavor attribute of aged cheese associated with the peptide fraction, but excessive levels are a defect leading to consumer rejection. Bitterness in cheese has been primarily associated with peptides that arise from the breakdown of casein. The last review of bitter peptides was published in 1992. This updated review compiled information about the bitter peptides published up to 2022. Our comprehensive search of the literature compiled 226 peptides associated with bitterness and cheese protein origins into a database (Supplemental Materials). The influences of a peptide's physical properties, such as molecular weight, average hydrophobicity, peptide length, number of prolines and the presence of hydrophobic amino acids in the peptide's terminus, were assessed for correlation with bitterness threshold values this assessment found that, among variables considered, higher molecular weight had the strongest correlation with higher bitterness among known peptides. Heatmaps of bitter peptides and their bitterness threshold values highlight ß-casein as the primary source of known bitter peptides in cheese. This comprehensive database of cheese protein-derived bitter peptides and this discovery of the correlation of a peptide's physical properties to bitterness will aid future researchers in the identification and discovery of contributors to cheese bitterness.

Comparison of milk oligosaccharides between goats with and without the genetic ability to synthesize αs1-casein.

Milk oligosaccharides (OS)-free complex carbohydrates-confer unique health benefits to the nursing neonate. Though human digestive enzymes cannot degrade these sugars, they provide nourishment to specific commensal microbes and act as decoys to prevent the adhesion of pathogenic micro-organisms to gastrointestinal cells. At present, the limited quantities of human milk oligosaccharides (HMO) impede research on these molecules and their potential applications in functional food formulations. Considerable progress has been made in the study of OS structures; however, the synthetic pathways leading to their synthesis in the mammary gland are poorly understood. Recent studies show that complex OS with fucose and N-acetyl neuraminic acid (key structural elements of HMO bioactivity) exist in goat milk. Polymorphisms in the CSN1S1 locus, which is responsible for synthesis of αs1-casein, affect lipid and casein micelle structure in goat milk. The present study sought to determine whether CSN1S1 polymorphisms also influence goat milk oligosaccharide (GMO) production and secretion. The GMO compositions of thirty-two goat milk samples, half of which were from genotype A/A (αs1-casein producers) and half from genotype O/O (αs1-casein non-producers), were determined with nanoflow liquid chromatography high-accuracy mass spectrometry. This study represents the most exhaustive characterization of GMO to date. A systematic and comprehensive GMO library was created, consolidating information available in the literature with the new findings. Nearly 30 GMO, 11 of which were novel, were confirmed via tandem mass spectrometric analyses. Six fucosylated OS were identified; 4 of these matched HMO compositions and three were identified for the first time in goat milk. Importantly, multivariate statistical analysis demonstrated that the OS profiles of the A/A and O/O genotype milks could be discriminated by the fucosylated OS. Quantitative analysis revealed that the goat milk samples contained 1.17 g/L of OS; however, their concentration in milks from A/A and O/O genotypes was not different. This study provides evidence of a genetic influence on specific OS biosynthesis but not total OS production. The presence of fucosylated GMO suggests that goat milk represents a potential source of bioactive milk OS suitable as a functional food ingredient.