Search Results (25)

β-Casein A1 and A2 (β-CN-A1 and β-CN-A2) are the two predominant β-CN proteoforms in bovine milk. β-CN-A1 has been associated with a greater propensity to release opioid peptides, such as β-casomorphin-7 (β-CM-7) and β-casomorphin-5 (β-CM-5), during gastrointestinal (GI) digestion, which may have adverse biological effects. This has stimulated growing interest in milk from cows carrying the β-CN A2A2 genotype (A2 milk), which requires reliable characterization methods. In this work, we developed a rapid, selective, and sensitive capillary electrophoresis–mass spectrometry (CE-MS) method for the accurate identification and quantification of β-CM-7 and β-CM-5 in milk hydrolysates from in vitro GI digestion of bovine milk. The method showed good linearity (R² > 0.99, over 0.5–100 mg/L for β-CM-7 and 0.25–100 mg/L for β-CM-5), limits of detection (0.25 and 0.10 mg/L), and repeatability (<0.2% for times and <1.4% for areas), and tandem mass spectrometry (MS/MS) allowed confirmation. The method was applied to A1A1 and A2A2 milk digested using the standardized INFOGEST protocol, followed by solid-phase extraction. β-CM-7 was detected and quantified only in A1A1 digests (0.98 mg/L), whereas β-CM-5 was not detected (<0.10 mg/L). These results indicate a differential release of β-CMs from A1 and A2 milk and support the method’s suitability for β-CM profiling, which may help assess A2 milk quality control and β-CM health impact. Full article

Background/Objectives: Endothelial dysfunction plays a central role in the development of cardiovascular diseases. β-Casomorphin-7 (BCM-7), a biologically active peptide generated during the digestion of A1 β-casein, is presumed to contribute to this process; however, its direct effects on endothelial cells have not been previously investigated. Here, we aimed to assess whether BCM-7 treatment induces endothelial cell dysfunction through inflammatory cytokines and reactive oxygen species (ROS). Methods: In our study, we analyzed the effects of BCM-7 (5 µg/mL) in combination with lipopolysaccharide (LPS, 100 ng/mL) on human umbilical vein endothelial cells (HUVECs/TERT2). The cell viability, apoptosis, necrosis, and intracellular reactive oxygen species were measured. Furthermore, proinflammatory cytokines and enzymes involved in the regulation of inflammation were assessed with quantitative real-time PCR. The gene and protein expression of enzymes that regulate inflammation and vascular function, thus maintaining endothelial homeostasis were assessed. Results: BCM-7 enhanced intracellular ROS production p ≤ 0.001, increased the expression of interleukin-6 (IL-6) and interleukin-8 (IL-8) p ≤ 0.001, and was more effective when used in combination with LPS p ≤ 0.001. It decreased the expression of cyclooxygenase-1 (COX-1) p ≤ 0.05, during 4 h of exposure, whereas it increased the expression of cyclooxygenase-2 (COX-2) p ≤ 0.001, lipoxygenase-5 (LOX-5) p ≤ 0.01, and nitric oxide synthase 3 (NOS3) p ≤ 0.001; prostaglandin D2 synthase (PTGDS) (p ≤ 0.05), expression was also increased after short treatment. Conclusions: Our results suggest that BCM-7 may contribute to the development of endothelial dysfunction, especially in the presence of LPS, by enhancing oxidative stress and inflammatory response. Full article

Background: β-casomorphin-10 (CM-10), a peptide fragment derived from milk casein with the sequence YPFPGPIPNS, has demonstrated notable anxiolytic activity in BALB/c mice. Yet, its cellular responses and mechanistic pathways remain largely uncharacterized. Methods: We performed RNA-seq analysis to profile gene expression changes in δ-opioid receptor-expressing HEK293 cells (DOR-HEK), comparing CM-10-treated and untreated conditions. Results: CM-10 exposure led to differential expression of 1714 genes in DOR-HEK cells, with 34 upregulated (>1.4-fold) (1.9%) and 1680 downregulated (<0.71-fold) (98.1%), based on a predicted p-value threshold of <0.05. Notably, we identified 10 clusters that were associated with reduced cyclic AMP (cAMP) in DOR-HEK cells following CM-10 treatment. These clusters particularly involved genes related to regulatory subunits of cAMP-dependent protein kinases, such as PRKAR2A, cAMP-responsive element-binding pathway, circadian rhythms, such as CLOCK, ARNT1, CRY2, PER1, and PER2, and anxiety and depression, such as NOTCH1, NOTCH2 and ANK2. A network with these selected genes was confirmed by STRING analysis. Conclusions: These findings indicate that CM-10 may activate DOR-mediated signaling by suppressing cAMP levels, implicating a distinct molecular cascade in HEK293 cells. Full article

The A1 and A2 variants of bovine β-casein (CSN2) have gained attention in the dairy industry due to potential health effects. The A1 variant, prevalent in Holstein-Friesian cattle, is a major source of β-casomorphin-7 (BCM-7)—an opioid-like peptide released during digestion and associated with lower digestive comfort. In this study, β-casein A1 and A2 variants and BCM-7 levels were quantified in raw milk and three commonly consumed dairy products (pasteurized milk, UHT milk, and milk powder) using ELISA. The samples came from dairy plants within a single operating zone. The A1 variant was significantly more frequent (13.69–22.41 ng/mL) than the A2 variant (8.10–12.60 ng/mL), although the local Holstein cattle population had a higher frequency of the A2 allele (63%) than A1 (37%). This discrepancy could be due to a more efficient expression of the A1 allele in cows with heterozygous or A1A1 genotypes. BCM-7 levels were low and did not vary significantly with CSN2 genotype or processing method. These results provide new insights into the composition of dairy products and contribute to the ongoing debate on the health implications and consumer acceptance of milk with the A1 β-casein variant. Full article

The digestion of A1 β-casein present in conventional milk releases β-casomorphin-7 (βCM-7), a bioactive peptide with potential implications for gastrointestinal and neurological health. A scoping review was performed to respond to the following research question: What are the health effects of consuming milk containing the A1 β-casein variant compared to the exclusive consumption of the A2 variant in humans? The evidence collected in this review of human studies with different populations (i.e., children, middle-aged adults, athletes) suggests that the consumption of milk containing A1 β-casein may negatively influence gut health by altering microbial composition, reducing intestinal motility, and increasing colonic fermentation, leading to elevated gas production and altered short-chain fatty acid (SCFA) profiles. The release of βCM-7 upon digestion can also compromise intestinal-barrier integrity, which may exacerbate symptoms of lactose intolerance, irritable bowel syndrome (IBS), or other allergy-related sensitivities. Its ability to cross the blood–brain barrier raises concerns about potential neurological effects. In contrast, milk containing exclusively A2 β-casein is associated with improved gastrointestinal outcomes, including the enhanced abundance of beneficial bacteria such as Bifidobacterium spp. and reduced inflammatory markers. Full article

Cataract formation remains a significant cause of global visual impairment. Increasing attention has been directed toward antioxidant-based interventions as potential non-surgical strategies to delay or prevent cataractogenesis, particularly in the age-related and diabetic contexts. This review summarizes recent preclinical evidence on nutritional antioxidants for the prevention of age-related and diabetic cataracts. Agents such as trimetazidine, Moringa oleifera stem extract, ginsenoside Rg1, lanosterol nanoparticles, β-casomorphin-7, and cerium oxide-based nanotherapies have been shown to mitigate oxidative damage, modulate redox signaling pathways, and preserve lens clarity. Advances in drug delivery, including topical formulations, nanoparticle carriers, and intravitreal injections, have been proposed to overcome the anatomical and pharmacokinetic barriers associated with the avascular lens. The new data support ongoing translational research to maximize the clinical use of antioxidants and highlight their therapeutic potential in the prevention of age-related and diabetic cataracts. Full article

Cow’s milk contains A1- and A2-β-caseins. The breakdown of A1-β-casein produces β-casomorphin-7 (BCM-7), a peptide with opioid-like properties that is associated with health aspects. In addition, A1- and A2-β-casein have different technological properties. The aim of the present study was to investigate whether cheese produced from the milk of homozygous A1A1 and A2A2 cows varies in terms of its physicochemical parameters and BCM-7 concentration. These parameters were analyzed during initial cheese processing, six weeks of ripening and 84 days of storage, including additional microbiological analyses during the storage period. The pH values of the A1A1 cheeses were higher than those of the A2A2 cheeses from the beginning of production until the starter culture bacteria were added. The yellowness values of the A1A1 cheeses were lower until the salt bath treatment. Water activity, lightness, hardness, fat, protein, NaCl and dry matter content, as well as color and microbiological parameters, were not affected by the β-casein genotype. BCM-7 concentrations were higher in the A1A1 cheeses after pressing and during ripening. We found mainly comparable quality characteristics and slightly different BCM-7 levels in the A1A1 and A2A2 cheeses. From this point of view, both varieties are equally suitable for cheese production. Full article

β-Casomorphin-7 (BCM), a breakdown product of milk β-casein, exhibits opioid activity. Opioids are known to affect the immune system, but the effects of BCM on ulcerative colitis (UC) are not clear. We examined the effects of BCM on mucosal immunity using a mouse dextran sulfate sodium-induced colitis model and an in vitro CD8+ T cell activation model. Human UC patients were examined to reveal the relationship between CD10 and mucosal immunity. Combined treatment of the colitis model with thiorphan (TOP) inhibited BCM degradation by suppressing CD10 in the intestinal mucosa, activating mouse mucosal CD8, and suppressing CD4 and Treg. In the CD8+ T cell in vitro activation assay using mouse splenocytes, BCM inhibited the oxidative phosphorylation (OXPHOS) of CD8+ T cells and induced the glycolytic pathway, promoting their activation. Conversely, in a culture system, BCM suppressed OXPHOS and decreased defensin α production in IEC6 mouse intestinal epithelial cells. In the mouse model, BCM reduced defensin α and butyrate levels in the colonic mucosa. During the active phase of human ulcerative colitis, the downward regulation of ileal CD10 expression by CpG methylation of the gene promoter was observed, resulting in increased CD8 activation and decreased defensin α and butyrate levels. BCM is a potential aggravating factor for UC and should be considered in the design of dietary therapy. In addition, decreased CD10 expression may serve as an indicator of UC activity and recurrence, but further clinical studies are needed. Full article

Special attention is given to cow’s milk and its variants, with ongoing discussions about health-related impacts primarily focusing on the A1 variant in contrast to the A2 variant. The difference between these variants lies in a single amino acid alteration at position 67 of β-casein. This alteration is presumed to make the A1 variant more susceptible to enzymatic breakdown during milk digestion, leading to an increased release of the peptide β-casomorphin-7 (BCM-7). BCM-7 is hypothesized to interact with µ-opioid receptors on immune cells in humans. Although BCM-7 has demonstrated both immunosuppressive and inflammatory effects, its direct impact on the immune system remains unclear. Thus, we examined the influence of A1 and A2 milk on Concanavalin A (ConA)-stimulated human peripheral blood mononuclear cells (PBMCs), as well as the effect of experimentally digested A1 and A2 milk, containing different amounts of free BCM-7 from β-casein cleavage. Additionally, we evaluated the effects of pure BCM-7 on the proliferation of ConA-stimulated PBMCs and purified CD4⁺ T cells. Milk fundamentally inhibited PBMC proliferation, independent of the β-casein variant. In contrast, experimentally digested milk of both variants and pure BCM-7 showed no influence on the proliferation of PBMCs or isolated CD4⁺ T cells. Our results indicate that milk exerts an anti-inflammatory effect on PBMCs, regardless of the A1 or A2 β-casein variant, which is nullified after in vitro digestion. Consequently, we deem BCM-7 unsuitable as a biomarker for food-induced inflammation. Full article

Bovine milk is an essential supplement due to its rich energy- and nutrient-rich qualities. Caseins constitute the vast majority of the proteins in milk. Among these, β-casein comprises around 37% of all caseins, and it is an important type of casein with several different variants. The A1 and A2 variants of β-casein are the most researched genotypes due to the changes in their composition. It is accepted that the A2 variant is ancestral, while a point mutation in the 67th amino acid created the A1 variant. The digestion derived of both A1 and A2 milk is BCM-7. Digestion of A2 milk in the human intestine also forms BCM-9 peptide molecule. The opioid-like characteristics of BCM-7 are highlighted for their potential triggering effect on several diseases. Most research has been focused on gastrointestinal-related diseases; however other metabolic and nervous system-based diseases are also potentially triggered. By manipulating the mechanisms of these diseases, BCM-7 can induce certain situations, such as conformational changes, reduction in protein activity, and the creation of undesired activity in the biological system. Furthermore, the genotype of casein can also play a role in bone health, such as altering fracture rates, and calcium contents can change the characteristics of dietary products. The context between opioid molecules and BCM-7 points to a potential triggering mechanism for the central nervous system and other metabolic diseases discussed. Full article

β-casomorphin 7 (BCM7) is a bioactive peptide that is released during the digestion of the β-casein (in particular, the A1 variant) present in cow’s milk. BCM7 has been linked to several health concerns such as gastrointestinal disorders. Milk processing alters the composition of milk, which in turn may affect its digestion, thus impacting the amount of BCM7 that is released. This study aimed to understand the impact of microfiltration on BCM7 release after the in vitro digestion (mimicking in vivo digestion) of semi-skimmed filtered milk compared to that of pasteurized milk and pasteurized Jersey milk (which does not contain A1 β-casein, the main source of BCM7). LC/MS was used to quantify BCM7. The results indicated that the β-casein variants present in milk, rather than the milk treatments themselves, are the key factors for the release of BCM7. Similar BCM7 levels were found in the filtered and pasteurized milk samples, whereas the Jersey milk released just half the amount. Full article

β-Casomorphin-7 (BCM-7) is a peptide released through the proteolysis of β-casein (β-CN), which is considered a bioactive peptide displaying evidence of promoting the binding and activation of the μ-opioid receptor located in various body parts, such as the gastrointestinal tract, the immune system and potentially the central nervous system. The possible effects of BCM-7 on health are a theme rising in popularity due to evidence found in several studies on the modulation of gastrointestinal proinflammatory responses that can trigger digestive symptoms, such as abdominal discomfort. With the advancement of studies, the hypothesis that there is a correlation of the possible effects of BCM-7 with the microbiota–gut–brain axis has been established. However, some studies have suggested the possibility that these adverse effects are restricted to a portion of the population, and the topic is controversial due to the small number of in vivo studies, which makes it difficult to obtain more conclusive results. In addition, a threshold of exposure to BCM-7 has not yet been established to clarify the potential of this peptide to trigger physiological responses at gastrointestinal and systemic levels. The proportion of the population that can be considered more susceptible to the effects of BCM-7 are evidenced in the literature review. The challenges of establishing the adverse effects of BCM-7 are discussed, including the importance of quantifying the BCM-7 release in the different β-CN genotypes. In summary, the reviewed literature provides plausible indications of the hypothesis of a relationship between β-CN A1/BCM-7 and adverse health effects; however, there is need for further, especially in vivo studies, to better understand and confirm the physiological effects of this peptide. Full article

It is known that casein hydrolysis accelerates gastrointestinal transit in comparison to intact casein, although the effect of the protein hydrolysis on the composition of the digests is not fully understood. The aim of this work is to characterize, at the peptidome level, duodenal digests from pigs, as a model of human digestion, fed with micellar casein and a previously described casein hydrolysate. In addition, in parallel experiments, plasma amino acid levels were quantified. A slower transit of nitrogen to the duodenum was found when the animals received micellar casein. Duodenal digests from casein contained a wider range of peptide sizes and a higher number of peptides above five amino acids long in comparison with the digests from the hydrolysate. The peptide profile was markedly different, and although β-casomorphin-7 precursors were also found in hydrolysate samples, other opioid sequences were more abundant in the casein digests. Within the same substrate, the evolution of the peptide pattern at different time points showed minimal changes, suggesting that the protein degradation rate relies more on the gastrointestinal location than on digestion time. Higher plasma concentrations of methionine, valine, lysine and amino acid metabolites were found in animals fed with the hydrolysate at short times (<200 min). The duodenal peptide profiles were evaluated with discriminant analysis tools specific for peptidomics to identify sequence differences between both substrates that can be used for future human physiological and metabolic studies. Full article

For over 20 years, bovine beta-casein has been a subject of increasing scientific interest because its genetic A1 variant during gastrointestinal digestion releases opioid-like peptide β-casomorphin-7 (β-CM-7). Since β-CM-7 is involved in the dysregulation of many physiological processes, there is a growing discussion of whether the consumption of the β-casein A1 variant has an influence on human health. In the last decade, the number of papers dealing with this problem has substantially increased. The newest clinical studies on humans showed a negative effect of variant A1 on serum glutathione level, digestive well-being, cognitive performance score in children, and mood score in women. Scientific reports in this field can affect the policies of dairy cattle breeders and the milk industry, leading to the elimination of allele A1 in dairy cattle populations and promoting milk products based on milk from cows with the A2A2 genotype. More scientific proof, especially in well-designed clinical studies, is necessary to determine whether a little difference in the β-casein amino acid sequence negatively affects the health of milk consumers. Full article

Although milk consumption is increasing worldwide, in some geographical regions, its consumption has persistently declined in recent decades. This fact, together with the increase in milk production prices, has caused both milk producers and the dairy industry to be immersed in a major crisis. Some possible solutions to this problem are to get people who do not currently consume milk to start drinking it again, or to market milk and dairy products with a higher added value. In this context, a type of milk called A2 has recently received attention from the industry. This type of milk, characterized by a difference in an amino acid at position 67 of the β-casein polypeptide chain, releases much smaller amounts of bioactive opioid peptide β-casomorphin 7 upon digestion, which has been linked to harmful effects on human health. Additionally, A2 milk has been attributed worse technological properties in the production of some dairy products. Thus, doubts exist about the convenience for the dairy industry to bet on this product. The aim of this review is to provide an update on the effects on human health of A2 milk, as well as its different technological properties to produce dairy products. Full article