Absolute quantification of two antimicrobial peptides α S2 -casein 182–207 and α S2 -casein 151–181 in bovine milk by UHPLC–ESI–MS/MS in sMRM mode

Abstract To evaluate if the recently identified antimicrobial milk peptides α S2 -casein 182–207 and α S2 -casein 151–181 can be of physiological or technological relevance, their presence in commercial milk was investigated. Thus, a UHPLC–ESI–MS/MS scheduled multiple reaction monitoring method was developed for quantification using stable isotope-labeled peptides. The developed protocol gave very good linear response (R 2  > 0.99) for both peptides, recovery >95% and coefficients of variations of 5.51% and 4.79%. The limits of detection (0.3 nM α S2 -casein 182–207 and 0.5 nM α S2 -casein 151–181 ) were low enough to detect both peptides without further enrichment. The highest peptide concentration was recorded in pasteurized milk (1.36 μM α S2 -casein 182–207 and 0.20 μM α S2 -casein 151–181 ). The concentration of α S2 -casein 182–207 was in a range that was previously reported to act bacteriostatic against Bacillus subtilis . Industrial milk processing led to a significant loss of both peptides resulting in a mean concentration of 0.47 µM α S2 -casein 182–207 and 0.098 µM α S2 -casein 151–181 in ultrahigh-temperature milk.