Identification of β-casomorphins 3 to 7 in cheeses and in their in vitro gastrointestinal digestates

•The content of β-casomorphins (BCMs) 3 to 7 in 12 bovine milk cheeses was determined.•Changes in the BCMs level during in vitro gastrointestinal digestion were examined.•Only BCM4 and BCM7 were released in the gastric or intestinal digestates of cheeses.•BCM7 level of a cheese serving size was not compatible with the in vivo opioid activity.

The occurrence of the bovine β-casein-derived casomorphins YPF (f60-62, BCM3), YPFP (f60-63, BCM4), YPFPG (f60-64, BCM5), YPFPGP (f60-65, BCM6) and YPFPGPI (f60-66, BCM7) was investigated in different cheeses by Ultra Performance Liquid Chromatography/High-Resolution Mass Spectrometry (UPLC/HR-MS). A total BCMs content was revealed in the range 0-11.74 mg kg−1. The amount of BCM7 varied from 0 to 7.63 mg kg−1. Gorgonzola cheese presented the highest BCM7 quantity, and it was the only sample to contain all the searched BCMs. Cheddar, Gorgonzola, Maasdam and Grana Padano were assessed for BCMs content following a recently proposed standardized in vitro static gastrointestinal digestion method (Minekus et al., 2014). After the gastric step, BCM7 was found only in Cheddar and Gorgonzola (0.25 mg kg−1 and 2.87 mg kg−1, respectively), along with BCM4. Only the latter peptide was detected in Grana Padano, whereas none of the studied BCMs were identified in the digested Maasdam. At the end of intestinal phase, all samples contained BCM7 (0.12-1.26 mg kg−1); the BCM4 was released only in digested Gorgonzola. Based on these results, in vitro digestion of a serving size of the studied cheeses does not potentially release a BCM7 amount compatible with an in vivo opioid activity.