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This work aimed to study the opioid peptide β-casomorphin-7 (BCM7) degradation or stability during digestion using human gastrointestinal (GI) juices and porcine jejunal brush border membrane (BBM) peptidases. Synthetic BCM7 was subjected to in vitro digestion by GI fluids obtained from human volunteers for 180 min, and to downstream degradation with porcine BBM vesicles. The BCM7 was sampled at 4 time points over 24 h after BBM addition. The digests were profiled by HPLC-electrospray ionization mass spectrometry (ESI/MS) to monitor BCM7 during GI digestion, and intact BCM7 through BBM digestion was quantified by reverse-phase (RP)-HPLC. We found that BCM7 was partly digested with human GI enzymes, as 3 proteolytic fragments in addition to f(60-66) YPFPGPI were detected: f(62-66) FPGPI, f(60-65) YPFPGP and f(61-66) PFPGPI. The RP-HPLC analysis revealed that 42% of the initial peptide was degraded after only 2 h of BBM digestion, and as much as 79% was degraded after 4-h digestion with supplementation of BBM. In conclusion, this study showed that most of BCM7 was degraded during GI and BBM digestion, although a small amount (5%) was still detected after 24-h digestion. It remains to be studied whether the small amount of intact BCM7 detected after in vitro digestion is transported via active transceptors in the human intestinal epithelial cells and enters blood circulation.

Celiac disease (CeD) is an autoimmune enteropathy triggered by immunogenic gluten peptides released during the gastrointestinal digestion of wheat. Our aim was to identify T cell epitope-containing peptides after ex vivo digestion of ancestral (einkorn, spelt and emmer) and common (hexaploid) wheat (Fram, Bastian, Børsum and Mirakel) using human gastrointestinal juices. Wheat porridge was digested using a static ex vivo model. Peptides released after 240 min of digestion were analyzed by liquid chromatography coupled to high-resolution mass spectrometry (HPLC-ESI MS/MS). Ex vivo digestion released fewer T cell epitope-containing peptides from the ancestral wheat varieties (einkorn (n = 38), spelt (n = 45) and emmer (n = 68)) compared to the common wheat varieties (Fram (n = 72), Børsum (n = 99), Bastian (n = 155) and Mirakel (n = 144)). Neither the immunodominant 33mer and 25mer α-gliadin peptides, nor the 26mer γ-gliadin peptide, were found in any of the digested wheat types. In conclusion, human digestive juice was able to digest the 33mer and 25mer α-gliadin, and the 26mer γ-gliadin derived peptides, while their fragments still contained naive T cell reactive epitopes. Although ancestral wheat released fewer immunogenic peptides after human digestion ex vivo, they are still highly toxic to celiac patients. More general use of these ancient wheat variants may, nevertheless, reduce CeD incidence.

Safety is the most important criteria of any substance or microorganism applied in the food industry. The whole-genome sequencing (WGS) of an indigenous dairy isolate LL16 confirmed it to be Lactococcus lactis subsp. lactis with genome size 2,589,406 bp, 35.4% GC content, 246 subsystems, and 1 plasmid (repUS4). The Nextera XT library preparation kit was used to generate the DNA libraries, and the sequencing was carried out on an Illumina MiSeq platform. In silico analysis of L. lactis LL16 strain revealed non-pathogenicity and the absence of genes involved in transferable antimicrobial resistances, virulence, and formation of biogenic amines. One region in the L. lactis LL16 genome was identified as type III polyketide synthases (T3PKS) to produce putative bacteriocins lactococcin B, and enterolysin A. The probiotic and functional potential of L. lactis LL16 was investigated by the presence of genes involved in adhesion and colonization of the host’s intestines and tolerance to acid and bile, production of enzymes, amino acids, and B-group vitamins. Genes encoding the production of neurotransmitters serotonin and gamma-aminobutyric acid (GABA) were detected; however, L. lactis LL16 was able to produce only GABA during milk fermentation. These findings demonstrate a variety of positive features that support the use of L. lactis LL16 in the dairy sector as a functional strain with probiotic and GABA-producing properties.