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.

Worldwide research on the health effects of bovine milk containing A1 and A2 ß-casein: Unraveling the current scenario and future trends through bibliometrics and text mining.

The possible adverse effect of consuming bovine milk with A1 ß-casein (but not with A2 ß-casein) on health aspects due to the release of ß-casomorphin-7 (BCM-7) is currently under debate. The aim of this study was to perform a bibliometric analysis of studies extracted from Scopus to explore the relationship between BCM-7, A1 or A2 bovine milk with different aspects of health. Over time, several research groups were formed that are no longer active and although some authors have returned to the field of study, they have focused their efforts mainly on conducting reviews that show the same imprecise conclusions due to the few original articles. Research is concentrated in Europe and Asia, where New Zealand, China and Germany are the countries with the most publications, records and citations on the subject, respectively. On the other hand, no country in Africa or South America has scientific production, which opens the possibility of building collaborations between countries and exploring areas that lack scientific studies. Based on conflicting information from primarily in vitro and animal studies, and limited clinical trials with poor designs, A1 milk presents pro-inflammatory and oxidative activity, but the evidence is insufficient to associate its consumption with negative health effects. However, A2 milk may be better tolerated by the digestive system of some individuals, suggesting its possible modulating role in the intestinal microbiota. Stronger scientific evidence is needed to reach a consensus on whether the presence of ß-casein A1 can significantly negatively affect health. The information shown will allow a better understanding of the subject and consumers will be able to make their own decisions regarding A1 or A2 milk.

A highly sensitive PCR method for A1 allele detection in A2 milk samples without DNA isolation.

We previously developed a genotyping method to detect the A1 and A2 alleles of the bovine ß-casein gene. This method required DNA extraction from hair samples. Recently, demand for A2 milk (milk from cows homozygous for the A2 allele) has increased, and dairy farms are required to have certification to produce A2 milk. Here, we describe the development of a new, simple, and sensitive genotyping method for the ß-casein gene that does not require DNA extraction. This method uses the CycleavePCR technique and can amplify the ß-casein gene directly from raw milk samples. Genotypes obtained from the milk samples (n = 27) were completely coincident with those obtained from genomic DNA. In addition, this method could quantify the A1 allele in the milk samples. The limit of detection for the A1 allele in A2 milk was 2%. The copy numbers of the A1 allele corresponding to the 2% detection limit were estimated to be 30.5 ± 24.3 molecules/µL. These findings indicate that this new genotyping method is simple and fast for detecting the A1 allele in milk samples and can therefore be potentially used to certify A2 milk.

Potential status of A1 and A2 variants of bovine beta-casein gene in milk samples of Indian cattle breeds.

This study aimed to determine the polymorphism in 7th exon of beta-casein gene (CSN2) gene in seven domestic (Kosali, Tharparkar, Gangatiri, Sahiwal, Gir, Khariar, Motu) and two exotic cattle breeds (Jersey and Holstein-Friesian). Genomic DNA was extracted from 1000 milk samples, and the C > A polymorphism in CSN2 was determined using the tetra-primer amplification refractory mutation system-polymerase chain reaction method. In all Indigenous cattle breeds, the mean frequency of A1A2 and A2A2 genotypes was 0.19 and 0.80, respectively. The A1A1 genotype was absent in all seven domestic cattle breeds. The frequency of the A2A2 genotype was highest in the Gir breed (0.93). However, the Sahiwal, Tharparkar, and Motu breeds also had a higher frequency of A2A2 genotype compared to other breeds. In contrast, Gangatiri breed of India showed lowest frequency of A2A2 genotype. The mean A1 and A2 allele frequency was 0.09 and 0.91, respectively. In exotic breeds, the mean frequencies of the A1A1, A1A2, and A2A2 genotypes were 0.42, 0.55, and 0.03, respectively. Similarly, the mean A1 and A2 allele frequency was 0.69 and 0.31, respectively. This study suggests the high potential of Gir, Sahiwal, Tharparkar, and Motu cattle for A2 milk production since they carry a favorable A2 genotype.

Probing of alpha, beta and kappa-caseins polymorphic variants in Gangatiri cow milk with the use of polyacrylamide gel electrophoresis and HRAMS.

This research communication aimed to probe the genetic polymorphisms of alpha, beta, and kappa caseins in Gangatiri cows (an indigenous Indian cattle). Detection of variants has received considerable research interest in the dairy industry and animal breeding in recent years as a source of good quality protein, but also of bioactive peptides that may be linked to health implications. The polymorphic nature of casein fractions and their association with milk production traits, composition, and quality also attracted several efforts in evaluating the allelic distribution of different casein locus as a potential dairy trait marker. Molecular techniques of polyacrylamide gel electrophoresis and high-resolution accurate mass-spectrometry have been applied to this probe. Sequence analysis of different casein types in the cows showed the presence of four specific variants.

Proteolysis of Micellar ß-Casein by Trypsin: Secondary Structure Characterization and Kinetic Modeling at Different Enzyme Concentrations.

Tryptic proteolysis of protein micelles was studied using ß-casein (ß-CN) as an example. Hydrolysis of specific peptide bonds in ß-CN leads to the degradation and rearrangement of the original micelles and the formation of new nanoparticles from their fragments. Samples of these nanoparticles dried on a mica surface were characterized by atomic force microscopy (AFM) when the proteolytic reaction had been stopped by tryptic inhibitor or by heating. The changes in the content of ß-sheets, α-helices, and hydrolysis products during proteolysis were estimated by using Fourier-transform infrared (FTIR) spectroscopy. In the current study, a simple kinetic model with three successive stages is proposed to predict the rearrangement of nanoparticles and the formation of proteolysis products, as well as changes in the secondary structure during proteolysis at various enzyme concentrations. The model determines for which steps the rate constants are proportional to the enzyme concentration, and in which intermediate nano-components the protein secondary structure is retained and in which it is reduced. The model predictions were in agreement with the FTIR results for tryptic hydrolysis of ß-CN at different concentrations of the enzyme.

Detection of A2A2 genotype of beta casein protein (CSN2) gene in local, exotic and cross bred cattle in Pakistan.

Genetic variants of bovine Beta-casein protein (CSN2) gene especially A1 and A2 are the most important variants in dairy cattle. A1 milk protein is considered as risk factor for different disease and milk intolerance which release Beta-Casomorphin-7 during digestion which is a bioactive opioid but not released from A2 milk protein. This opioid is responsible for several human health problems like Coronary Heart disease, type 1 diabetics, milk intolerance and other neurological disorders. In present study, 360 blood sample were collected from Lohani, Achai, jersey, Holstein Friesian, Achai x jersey, Friesian x Sahiwal and Sahiwal x Friesian from different region of Khyber Pakhtunkhwa (KP) province. The polymerase chain reaction (PCR) amplicons were sequenced for the identification of polymorphism in exon 7 of Beta-casein protein (CSN2) gene. Sequencing analysis explored CSN2 genotype in exon 7 using the Genomic sequence from GenBank (X.71104) g.8101 C > A at codon 67. The allelic and genotypic frequencies of CSN2 gene were analyzed and observed that Holstein Friesian cattle exhibited A1A2 33%, A1A1 50% and A2A2 17%, Jersey cattle show 68% A1A1, 18% A1A2 and 14% A2A2, Sahiwal x Friesian 56% A1A1, 26% A1A2 and 18% A2A2, Jersey × Achai 78% A2A2, 15% A1A2 and 7% A1A1, Achai 100% A2A2 Lohani 100% A2A2. This is a preliminary study, conducted with meager resources, therefore, it is very difficult to make conclusion that which particular breed possess harmful alleles and which breed possess useful alleles of beta-casein gene. Therefore, a comprehensive molecular work is needed to be performed with greater number of samples sequencing.

Assessment of A1 and A2 variants in the CNS2 gene of some cattle breeds by using ACRS-PCR method.

The aim of this study is to reveal ß-casein polymorphism of some cattle breeds and also the potential to produce A2 milk from existing animals and to develop strategies in this area. Therefore, a total of 400 cattle, 100 animals from each breed of Holstein, Brown Swiss, Jersey and Simmental raised commonly in Turkey, were obtained, and C > A polymorphism in 67th amino acid in the 7th exons of ß-casein gene was determined by TaqI enzyme with PCR-ACRS method. Blood samples were collected from dairy cattle farms raising Holstein, Brown Swiss and Jersey breeds from Konya province and Simmental breed from Kütahya province in Turkey. A1 and A2 allele frequencies in Holstein, Brown Swiss, Jersey and Simmental cattle breeds were determined as 0.475/0.525, 0.370/0.630, 0.215/0.785 and 0.440/0.560, respectively. The highest A2 allele frequency (0.785) was found in Jersey breed. A1A1 genotypes in Holstein, Brown Swiss, Jersey and Simmental breeds were 0.240, 0.150, 0.030 and 0.160, respectively; A1A2 genotypes were 0.470, 0.440, 0.370 and 0.560, respectively; A2A2 genotypes were determined as 0.290, 0.410, 0.600 and 0.280, respectively. In these breeds, the highest A2A2 genotype frequency was found in Jersey (0.600), the lowest A1A1 genotype frequency (0.030) was found in Jersey and the highest A1A2 genotype frequency (0.560) was found in Simmental. Holstein, Brown Swiss, Simmental and Jersey populations were at the level of Hardy-Weinberg in terms of ß-casein gene (p > 0.05). The average Ho, He and PIC values were calculated as 0.460, 0.469 and 0.605, respectively. In conclusion, the frequency of commonly reared cattles in Turkey especially Brown Swiss, and Jersey breeds in A2A2 genotype are satisfactory, but it can be said that the use of animals with A2 allele in selection is extremely important for increasing A2 milk producing potential in the future.

The cysteine residue in beta-lactoglobulin reacts with oxidized tyrosine residues in beta-casein to give casein-lactoglobulin dimers.

Proteins are modified during milk processing and storage, with sidechain oxidation and crosslinking being major consequences. Despite the prevalence and importance of proteins in milk, and particularly caseins (∼80% of total content), the nature of the cross-links formed by oxidation, and their mechanisms of formation, are poorly characterized. In this study, we investigated the formation and stability of cross-links generated by the nucleophilic addition of Cys residues to quinones generated on oxidation of Tyr residues. The mechanisms and stability of these adducts was explored using ubiquitin as a model protein, and ß-casein. Ubiquitin and ß-casein were oxidized using a rose Bengal/visible light/O2 system, or by the enzyme tyrosinase. The oxidized proteins were incubated with glutathione or ß-lactoglobulin (non-oxidized, but unfolded by treatment at 70 °C), before analysis by SDS-PAGE, immunoblotting and LC-MS. Our data indicate that Cys-quinone adducts are readily-formed, and are stable for >48 h. Thus, oxidized ß-casein reacts efficiently with the thermally unfolded ß-lactoglobulin, likely via Michael addition of the exposed Cys to a Tyr-derived quinone. These data provide a novel, and possibly general, mechanism of protein cross-link formation, and provides information of the stability of these species that have potential as markers of protein quality.

Amyloid fibril formation by αS1- and ß-casein implies that fibril formation is a general property of casein proteins.

Caseins are a diverse family of intrinsically disordered proteins present in the milks of all mammals. A property common to two cow paralogues, αS2- and κ-casein, is their propensity in vitro to form amyloid fibrils, the highly ordered protein aggregates associated with many age-related, including neurological, diseases. In this study, we explored whether amyloid fibril-forming propensity is a general feature of casein proteins by examining the other cow caseins (αS1 and ß) as well as ß-caseins from camel and goat. Small-angle X-ray scattering measurements indicated that cow αS1- and ß-casein formed large spherical aggregates at neutral pH and 20°C. Upon incubation at 65°C, αS1- and ß-casein underwent conversion to amyloid fibrils over the course of ten days, as shown by thioflavin T binding, transmission electron microscopy, and X-ray fibre diffraction. At the lower temperature of 37°C where fibril formation was more limited, camel ß-casein exhibited a greater fibril-forming propensity than its cow or goat orthologues. Limited proteolysis of cow and camel ß-casein fibrils and analysis by mass spectrometry indicated a common amyloidogenic sequence in the proline, glutamine-rich, C-terminal region of ß-casein. These findings highlight the persistence of amyloidogenic sequences within caseins, which likely contribute to their functional, heterotypic self-assembly; in all mammalian milks, at least two caseins coalesce to form casein micelles, implying that caseins diversified partly to avoid dysfunctional amyloid fibril formation.

Worldwide Research Trends on Milk Containing Only A2 ß-Casein: A Bibliometric Study.

The protein fraction of ß-casein may play a key role in the manifestation of a new intolerance: milk protein intolerance. The most common forms of ß-casein among dairy cattle breeds are A1 and A2 ß-casein. During gastrointestinal digestion of A1 ß-casein, an opioid called peptide ß-casomorphin-7 (BCM-7) is more frequently released, which can lead to adverse health outcomes. For that reason, novel products labelled as "A2 milk" or "A1-free dairy products" have appeared on the market. In this context, a bibliometric analysis on A2 ß-casein research was carried out through the Web of Science (WoS) database. The main objective of this work was to provide an overview of the state of the art in the field of ß-casein A2 by analyzing the number of publications per year, trends in thematic content, the most frequently used terms, and the most important institutions and countries in the field. This bibliometric study showed that a greater effort is needed to determine the possible implications of this novel product for human health and the market.

Demographic pattern of A1/A2 beta casein variants indicates conservation of A2 type haplotype across native cattle breeds (Bos indicus) of India.

Genetic variations of the beta casein gene hold importance because of their probable association with human health. Comparative sequence analysis of ß-casein gene across Indian native, crossbred and exotic breeds in India revealed 15 SNPs and 4 INDELs corresponding to 14 haplotypes. The frequency of A2 type haplotype was maximum (0.941) across all Indian native breeds. Among the 15 variants reported for taurine breeds, only three (A1, A2 and B) were observed in analysed populations. Allelic profiling of A1/A2 ß-casein variants in ~ 4000 animals belonging to three cattle types and breeding bulls also revealed the predominance of A2 allele (0.95) in Indian cattle. The high proportion of A2 allele/haplotype indicates that Indian native cattle are the best suited to meet the demands for A2 milk globally. However, a higher percentage of heterozygous genotype (A1A2) in breeding bulls warrants the need to screen sire lines so as to drift the herd towards A2. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03232-0.

Molecular insights into the interaction of 5-fluorouracil and Fe3O4 nanoparticles with beta-casein: An experimental and theoretical study.

We investigated the potential carrier of milk beta-casein (ß-CN) and its interactions with 5-fluorouracil (5-FU) and iron oxide nanoparticles (Fe3O4 NPs). We used different spectroscopic methods of fluorescence, UV-Visble, circular dichroism (CD), synchronous fluorescence, zeta potential assay, and computational studies to clarify the protein interaction with 5-FU and Fe3O4 NPs. The fluorescence data indicated both Fe3O4 NPs and 5-FU could quench the intrinsic fluorescence of ß-CN. Fluorescence measurements showed that the single interaction of ß-CN with 5-FU or Fe3O4 NPs was static, while reacted ß-CN with both 5-FU and Fe3O4 NPs simultaneously showed a dynamic quenching. Synchronous fluorescence data in both tests revealed that the tryptophan (Trp) residue of ß-CN had a dominant role in quenching and the polarity of its microenvironment more than tyrosine (Tyr) increased in interaction with 5-FU. All the binding sites and thermodynamic parameters were obtained at 25, 37, and 42 °C. The analysis of thermodynamic parameters and Job's plot techniques pointed to that both of these complexes with the 1:1 M ratio were exothermic (ΔH°<0) driven with the van der Waals and H-bonding interactions (in agreement with the docking results). The CD spectra in the region of far-UV and thermal denaturation study indicated minor changes in the secondary structure of ß-CN in the presence of various concentrations of Fe3O4 NPs and 5-FU. Also, from the molecular dynamics (MD) analysis, as a result, the protein structure was stable during 100 ns. The outcomes highlighted that ß-CN protein could form a great bind with 5-FU and Fe3O4 NPs ligands (supporting the zeta potential assay results) by independent binding sites. These results would be helpful insight to construct a potential magnetic nanocarrier ß-CN base for 5-FU drug delivery.

Investigation of genetic polymorphism at ß-casein A1/A2 loci and association analysis with production & reproduction traits in Vrindavani crossbred cows.

Casein constitutes approximately 80% of the total protein in bovine milk and is regarded as a high-quality dietary protein embracing all the nine essential amino acids. However, the contested physiological effect of a bioactive peptide released upon digestion of a ß-casein milk protein variant originating from a cow of a particular genetic makeup has evoked wide interest in research and industry. Present investigations were carried out to genotype the polymorphism in milk ß-casein gene, delineate the seasonal, periodic, and parity variations in production and reproduction traits, and examine the genetic association between ß-casein genotypes and production, and reproduction traits in Vrindavani crossbred cows. The study revealed that all three types of genotypes viz. A1A1, A2A2 and A1A2 were present in the Vrindavani crossbred population with genotypic frequencies of 12.3%, 39.6% and 48.1% respectively. The least-squares analysis revealed that the season of calving, period of calving, and parity affected several production and reproduction traits of Vrindavani cows significantly. It was found that ß-Casein A1/A2 genotype had a significant effect on economic traits viz. LL (p ≤ 0.05), MY/LL (p ≤ 0.05), and Fat% (p ≤ 0.05) in Vrindavani crossbreds. The findings uncover the genetic constitution of the crossbreds for ß-casein locus and emphasize its relationship with important economic traits that can aid in devising selection goals.

A1- and A2 beta-casein on health-related outcomes: a scoping review of animal studies.

PURPOSE: Food-derived bioactive peptides may influence important physiological functions. An important example is beta-casomorphins, which are opioid peptides derived from A1 beta-casein in bovine milk and have been associated to be risk factors for non-communicable diseases in humans. A1 and A2 beta-casein are different with respect to the release of bioactive peptides, in particular BCM-7. However, evidence from human studies is limited and could be complemented with evidence derived from animal studies. We conducted a scoping review to identify animal studies investigating the effects of A1 beta-casein or BCM-7 compared to A2 beta-casein or any other intervention on health-related outcomes. METHODS: We systematically searched for relevant studies in two electronic databases (Medline, Embase; last search performed March 2020). Two reviewers independently undertook study selection and data extraction of included references. Results were summarized tabularly and narratively. RESULTS: We included 42 studies investigating various animal models, including rats, mice, rabbits, and dogs. Six studies investigated health-related outcomes of A1- vs. A2 milk, while most studies (n = 36) reported on physiological properties (e.g., analgesic effect) of BCM-7 as an opioid peptide. Included studies were extremely heterogeneous in terms of the study population, type of intervention and dose, and type of outcome measures. CONCLUSIONS: Only a few studies comparing the effects of A1- and A2 milk were identified. More studies addressing this research question in animal models are needed to provide essential information to inform research gaps. Results from future studies could eventually complement research for humans, particularly when the body of evidence remains uncertain as is the case in the A1- and A2 milk debate.

The Moo'D Study: protocol for a randomised controlled trial of A2 beta-casein only versus conventional dairy products in women with low mood.

BACKGROUND: Beta-casein is a major protein in cow's milk, of which A1 and A2 are the most frequent variants. Recent evidence implicates A1 beta-casein consumption in mechanisms that are of potential importance to mental health, yet its possible effects on psychological endpoints remains unknown. The primary aim of the study is to evaluate the comparative effects of consumption of dairy products containing A2 beta-casein versus conventional dairy (i.e. containing both A1 and A2 beta-casein) on symptoms of psychological distress in women with low mood. METHODS: 'The Moo'D Study' is a 16-week, superiority, 1:1 parallel group, triple-blinded, randomised controlled trial. Ninety women with low mood (Patient Health Questionnaire score ≥ 5) will be randomised to consume either A2 beta-casein only or conventional dairy products. The primary outcome, symptoms of psychological distress, will be measured by the 21-item Depression, Anxiety and Stress Scale. Secondary outcomes will include symptoms of depression, anxiety and stress, severity of low mood, cognition, gut microbiota composition, gut symptomatology, markers of immune function, gut inflammation, systemic metabolites, endothelial integrity and oxidative stress, body composition, perceived wellbeing, sleep, quality of life, resource use and cost-effectiveness. DISCUSSION: This study will advance our understanding of the possible impact of milk proteins on psychological distress in women as well as elucidate mechanisms underpinning any association. Given dairy products form a substantial component of traditional and Western diets, the implications of these findings are likely to be of clinical and public health importance. TRIAL REGISTRATION: The trial protocol has been prospectively registered with the Australia and New Zealand Clinical Trials Registry, ACTRN12618002023235 . Registered on 17 December 2018.

Estructura, propiedades y genética de las caseínas de la leche: una revisión / Structure, properties and genetic of milk caseins: a review / Estrutura, propriedades e genética das caseínas do leite: uma revisão

Resumen La leche es un alimento esencial para los humanos y una de sus importancias radica en el contenido de proteínas lácteas. Las proteínas más frecuentes en este preciado líquido son las caseínas (αS1-caseína, αS2-caseína, β-caseína y κ-caseína), las cuales son fuente de aminoácidos para la dieta de los mamíferos en sus primeros días de vida. En la leche, las caseínas, están formadas por agregados moleculares de proteínas de tamaños variables denominados micelas. El objetivo de esta revisión es presentar un panorama general de la estructura, propiedades y genética de las caseínas lácteas y su relación con la salud humana. A partir de esta revisión, se pudo establecer, que las αs1 y αs2 caseínas se encuentran en conjunto con la β-caseína, formando el núcleo micelar, interactuando con los iones de calcio, para formar y mantener la micela estable. Animales caracterizados genéticamente con algunas variantes de estas proteínas, se asocian con un rendimiento en el volumen de leche. La κ-caseína, por su parte, está asociada con un aumento en el rendimiento y calidad de los quesos, de ahí su importancia económica, mientras que las formas más comunes de β-caseína en razas de ganado lechero son A1 y A2. La β-caseína A2 no presenta efectos negativos a la salud humana, por el contrario, ha sido asociada con propiedades reductoras de colesterol y triacilglicéridos. Sin embargo, la variante A1 de la β-caseína produce un péptido bioactivo denominado β-casomorfina-7 (BCM-7), que puede desempeñar un papel etiológico poco claro en el desarrollo de algunas enfermedades en humanos, tales como: enfermedad isquémica del corazón, diabetes mellitus tipo 1, síndrome de muerte súbita infantil (SIDS), desórdenes neurológicos, como autismo y esquizofrenia.

Abstract Milk is an essential food for humans and one of the reasons of its importance lies in the content of milk proteins. The most frequent proteins in this precious liquid are caseins (αS1-casein, αS2-casein, β-casein and κ-casein), which are a source of amino acids for the diet of mammals in their first days of life. In milk, caseins are made up of molecular aggregates of proteins of varying sizes called micelles. The objective of this review is to present an overview of the structure, properties and genetics of dairy caseins and their relation with human health. From this review, it was established that αs1 and αs2 caseins are found together with β-casein, forming the micellar nucleus and interacting with calcium ions, to form and maintain stable the micelle. Animals genetically characterized with some variants of these proteins are associated with a yield in milk volume. For its part, κ-casein is associated with an increase in the yield and quality of cheeses, hence its economic importance, while the most common forms of β-casein in dairy cattle are A1 and A2. β-casein A2 does not have negative effects on human health; on the contrary, it has been associated with lowering properties of cholesterol and triacylglycerides. However, the A1 variant of β-casein produces a bioactive peptide called β-casomorphin-7 (BCM-7), which may play an unclear etiological role in the development of some diseases in humans, such as: ischemic heart disease, type 1 diabetes mellitus, sudden infant death syndrome (SIDS), neurological disorders, such as autism and schizophrenia.

Resumo O leite é um alimento essencial para o ser humano e uma de suas principais característica é o teor de proteínas do leite. As proteínas mais frequentes neste líquido são as caseínas (αS1-caseína, αS2-caseína, β-caseína e κ-caseína), que são uma fonte de aminoácidos para a dieta dos mamíferos nos primeiros dias de vida. As caseínas no leite são constituídas por agregados moleculares de proteínas de variados tamanhos, chamados micelas. O objetivo desta revisão é apresentar uma visão geral da estrutura, propriedades e genética das caseínas lácteas e sua relação com a saúde humana. A partir desta revisão, foi estabelecido que as caseínas αs1 e αs2 são encontradas em conjunto com a β-caseína, formando o núcleo micelar, interagindo com os íons cálcio, para formar e manter a micela estável. Animais geneticamente caracterizados com algumas variantes dessas proteínas estão associados com o rendimento da produção de leite. Por sua vez, a κ-caseína está associada ao aumento do rendimento da produção e da qualidade dos queijos, por isso sua importância econômica, enquanto as formas mais comuns de β-caseína em bovinos leiteiros são A1 e A2. A Β-caseína A2 não tem efeitos negativos na saúde humana, pelo contrário, tem sido associada a propriedades redutoras do colesterol e dos triacilglicéridos. No entanto, a variante A1 da β-caseína produz um peptídeo bioativo denominado β-casomorfina-7 (BCM-7), que pode desempenhar uma função etiológico ainda desconhecida no desenvolvimento de algumas doenças em humanos, tais como: doença isquêmica do coração, diabetes mellitus tipo 1, síndrome da morte súbita infantil (SMSL), distúrbios neurológicos, como autismo e esquizofrenia.

Genetic polymorphism of bovine beta-casein gene in Japanese dairy farm herds.

The aim of this study was to investigate beta-casein polymorphism among 320 Japanese cows sampled from eight dairy farms. We used a newly-developed genotyping method that involved collecting DNA from hairs and a Cycleave polymerase chain reaction (PCR) assay to detect the A1, A2, and B variants. Results revealed the presence of five genotypes (A1A1, A2A2, A1A2, A1B, and A2B). We found that the most common genotype was A2A2 (0.42), followed by A1A2 (0.39) and A1A1 (0.11). The A1B and A2B genotypes were less frequent (<0.05). The frequencies of alleles A1, A2, and B were calculated to be 0.32, 0.64, and 0.04, respectively. Our study is the first to show the current status of beta-casein polymorphisms in Japanese dairy farms. Given the adverse effects of A1 beta-casein on human health, attempts have been made to develop herds consisting solely of A2A2 cows. Our study provides a reference for improving cow populations in Japanese dairy farms. The Cycleave PCR-based assay we developed here can be used for rapid and reliable genotyping of bovine beta-casein.

Casomorphins and Gliadorphins Have Diverse Systemic Effects Spanning Gut, Brain and Internal Organs.

Food-derived opioid peptides include digestive products derived from cereal and dairy diets. If these opioid peptides breach the intestinal barrier, typically linked to permeability and constrained biosynthesis of dipeptidyl peptidase-4 (DPP4), they can attach to opioid receptors. The widespread presence of opioid receptors spanning gut, brain, and internal organs is fundamental to the diverse and systemic effects of food-derived opioids, with effects being evidential across many health conditions. However, manifestation delays following low-intensity long-term exposure create major challenges for clinical trials. Accordingly, it has been easiest to demonstrate causal relationships in digestion-based research where some impacts occur rapidly. Within this environment, the role of the microbiome is evidential but challenging to further elucidate, with microbiome effects ranging across gut-condition indicators and modulators, and potentially as systemic causal factors. Elucidation requires a systemic framework that acknowledges that public-health effects of food-derived opioids are complex with varying genetic susceptibility and confounding factors, together with system-wide interactions and feedbacks. The specific role of the microbiome within this puzzle remains a medical frontier. The easiest albeit challenging nutritional strategy to modify risk is reduced intake of foods containing embedded opioids. In future, constituent modification within specific foods to reduce embedded opioids may become feasible.

Análisis genético de cinco polimorfismos de nucleótido simple de caseínas lácteas obtenidos con chips genómicos en ganado Holstein de Antioquia, Colombia / Genetic analysis of five single nucleotide polymorphisms of dairy caseins obtained with genomic chips in Holstein cattle from Antioquia, Colombia

RESUMEN Los polimorfismos genéticos asociados con las caseínas de la leche son de gran importancia, ya que pueden ser usados como marcadores genéticos para mejorar el rendimiento productivo en los hatos lecheros. El objetivo de este estudio fue evaluar la diversidad y estructura genética de 5 SNP de caseínas de la leche, obtenidos con chips genómicos en vacas y toros de raza Holstein en Antioquia (Colombia). Fueron muestreados 113 animales de raza Holstein en 3 regiones del departamento de Antioquia (norte, centro y oriente) y un cuarto grupo de sementales comerciales. Los animales fueron genotipificados con chips genómicos de alta densidad (Illumina BovineHD e Illumina SNP50 v2), a partir de los cuales se identificaron 5 SNP (ARS-BFGL-NGS-8140, BTA-77380-no-rs, BTA-32346-no-rs, BTB-00821654 y ARS-BFGL-NGS-15809). Para cada SNP se realizó un análisis genético mediante un análisis de varianza molecular (amova) usando el software GenAIEx 6.501. Los SNP con mayor heterocigosidad total (HT) fueron ARS-BFGL-NGS-8140 y BTA-32346-no-rs, con resultados cercanos al 45%; sin embargo, la Ht para ARS-BFGL-NGS-15809, BTA-77380-no-rs y BTB-00821654 estuvo por debajo del 15%. El SNP con mayor diversidad genética fue BTA-32346-no-rs (Ho-He = 0,06; p < 0,05). En esta investigación se evaluó una subpoblación de toros comerciales extranjeros, en la cual se obtuvieron frecuencias alélicas y genotípicas similares a las obtenidas para las subpoblaciones locales, sugiriendo que los alelos de los toros muy posiblemente están fijados en dichas subpoblaciones, por lo que la estructura y diversidad genética tienden a ser bajas en la muestra de estudio.

ABSTRACT Genetic polymorphisms associated with milk caseins have a great importance since they can be used as genetic markers to improve productive performance in dairy herds. The main goal of the present study was to evaluate the diversity and genetic structure of 5 SNPs of milk caseins, obtained with genomic chip in Holstein cows and bulls from Antioquia (Colombia). 113 Holstein animals were sampled in 3 regions of Antioquia (north, center, and east), and a fourth group of commercial sires. Animals were geno-typed with high-density SNP chips (Illumina BovineHD and Illumina SNP50 v2), from which 5 SNPs were identified (ARS-BFGL-NGS-8140, BTA-77380-no-rs, BTA-32346-no-rs, BTB-00821654 and ARS-BFGL-NGS-15809). For each SNP, a genetic analysis was performed by means of an analysis of molecular variance (AMOVA) using the GenAIEx 6.501 software. The SNPs with the highest total heterozygosity (Ht) were ARS-BFGL-NGS-8140 and BTA-32346-no-rs, with results close to 45%; however, the HT for ARS-BFGL-NGS-15809, BTA-77380-no-rs, and BTB-00821654 were below 15%. The SNP with the highest genetic diversity was BTA-32346-no-rs (Ho-He = 0,06; p < 0,05). In this research a subpopulation of foreign commercial bulls was evaluated, in which similar allelic and genotypic frequencies to those for local subpopulations were obtained, suggesting that the alleles of the bulls are very possibly fixed in these subpopulations, so that the structure and genetic diversity tend to be low in the study sample.