Milk Protein Casein Research Articles

Urinary opioid peptides in children with autism spectrum disorders. A Pilot Study

BACKGROUND: Circulating β-casomorphins (BCM) and gluten exorphins (GE), which are exogenous opioid peptides originating from bovine milk protein casein and cereal protein gluten, respectively, have been proposed as potential trigger factor to symptoms observed in children with autism. OBJECTIVE: Given the debate surrounding the detectability of these opioid peptides in body fluids, particularly using highly sensitive and specific mass spectrometry (HPLC-MS) techniques, we aimed to investigate their presence in urine samples of autistic subjects. METHODS: We employed an HPLC-MS method for peptide detection. RESULTS: The presence of several BCMs and GEs in the urine of both autistic children (ASD) and healthy controls (HC) was documented. The detection of dietary opioid peptides even at very low concentrations underscores the sensitivity of this novel HPLC-MS method. BCM-8 was more often detected in the ASD group compared to the HC group. A higher prevalence of gastrointestinal (GI) symptoms were also observed in the ASD group. CONCLUSIONS: This pilot study supports the presence of BCMs and GEs in urine samples in subjects with autism as well as healthy controls which was the main goal of this pilot study. Prolonged exposure to bovine BCMs and GEs may play a role in the manifestation of core and GI symptoms in subgroups of autism. Further research is warranted to investigate this phenomenon thoroughly.

Interaction of Sambiloto (Andrographis paniculata) Bioactive Compound with Milk Protein (Whey and Casein) Through Molecular Docking and Molecular Dynamics Simulation as a Basis for Encapsulation

This research aims for developing immune-boosting products necessary. The active ingredient in Andrographis paniculate (AP) acts as an immunostimulant which can improve the work of the immune system. The first stage of research was a collection of bioactive compounds from KnapSack database of Kanaya, Dr. Duke's Phytochemical and Ethnobotanical were compiled and selected based on the online pass results of each bioactive compound as an immunomodulator and 10 active compounds were obtained which will be continued. The second stage of research was a docking molecular between whey proteins (β-lactoglobulin and α-lactalbumin) with active compounds from AP and casein (α-Casein, β-Casein, and κ-Casein). The highest binding affinity was obtained for α-Casein with Neoandrographolide at -9.2 Kcal/mol. The results of the complex α-Casein with Neoandrographolide (CC) and α-Casein with Neoandrographolide ultraheat (CCT) support the research, namely to function α-Casein as an encapsulant well as a transporter or drug delivery of Neoandrographolide without changing the conformation of casein and disturbing its function. However, the conformation of casein will change drastically during ultraheat treatment to maintain the conformation and binding with the Neoandrographolide ligand. In addition, it supports the simulation results of single α-Casein at ultraheat temperatures which show conformational stability that is not much different from single α-casein and complexes at physiological temperatures

Synthesis and dual sensing of vanadium and copper ions using protein capped selenium nanoparticles

Highly homogenous and spherical shaped Selenium nanoparticles with an average size of 20 nm were synthesized from an eco-friendly and environmentally safe approach using bovine milk protein casein as stabilizing agent (named as CasSeNPs). Protein functionalization provided excellent stability at variable temperatures (25–60 °C), pH (8–10), ionic strength (0.05–2 M), and storage period for more than five months. Synthesized CasSeNPs were extensively characterized using various spectroscopic (UV–visible, Fourier transform infrared (FTIR), circular dichroism (CD), intrinsic fluorescence spectroscopy), particle sizing (dynamic light scattering (DLS) and zeta potential), microscopic (scanning electron microscopy, energy dispersive X-ray spectroscopy (SEM/EDS), atomic force microscopy (AFM)), and X-ray diffraction (XRD) measurements. Moreover, CasSeNPs was established as highly sensitive and selective sensor for the simultaneous detection of vanadium (V+2) and copper (Cu+2) ions with significant linearity (0.5–10 and 0.1–1.0 µM, respectively), detection limits (LOD = 0.03 and 0.067 µM, respectively), and percent recoveries (94–105 %) in real environmental (tap water) and biological (human plasma and urine) samples with adequate test performance. Cumulatively, obtained results revealed that the established very simple, biocompatible and economical nanosensor has great potential for applications in analytical chemistry as well as for biological investigations.

Basophil allergen threshold sensitivity to casein (casein-specific CD-sens) predicts allergic reactions at a milk challenge in most but not all patients.

The basophil activation test is an emerging clinical tool in the diagnosis of cow's milk allergy (CMA). The aim was to assess the association between the basophil allergen threshold sensitivity to the major milk protein casein (casein-specific CD-sens), the levels of milk- and casein-specific Immunoglobulin E antibodies (IgE-ab), and the severity of allergic reactions at milk challenges. We enrolled 34 patients aged 5-15 (median 9) years who underwent a double-blind placebo-controlled milk-challenge (DBPCMC) as screening before inclusion in an oral immunotherapy study for CMA. The severity of the allergic reaction at the DBPCMC was graded using Sampson's severity score. Venous blood was drawn before the DBPCMC. Milk- and casein-specific IgE-ab were analyzed. Following in vitro stimulation of basophils with casein, casein-specific CD-sens, was determined. Thirty-three patients completed the DBPCMC. There were strong correlations between casein-specific CD-sens and IgE-ab to milk (rs = 0.682, p < .001), and between casein-specific CD-sens and IgE-ab to casein (rs = 0.823, p < .001). There was a correlation between the severity of the allergic reaction and casein-specific CD-sens level (rs = 0.395, p = .041) and an inverse correlation between casein-specific CD-sens level and the cumulative dose of milk protein to which the patient reacted at the DBPCMC (rs = -0.418, p = .027). Among the 30 patients with an allergic reaction at the DBPCMC, 67% had positive casein-specific CD-sens, 23% had negative casein-specific CD-sens, and 10% were declared non-responders. Two thirds of those reacting at the DBPMC had positive casein-specific CD-sens, but reactions also occurred despite negative casein-specific CD-sens. The association between casein-specific CD-sens and the severity of the allergic reaction and cumulative dose of milk protein, respectively, was moderate.

Milk protein polymorphisms and casein haplotypes in Blanco Orejinegro cattle of Colombia

The aim was to determine the genetic variation in the CSN1S1, CSN2, CSN1S2, CSN3 and LGB genes in Blanco Orejinegro cattle. 419 animals from 15 herds were genotyped with GGP Bovine 150 K (n= 70) and 50 K (n= 349) chips. Information was obtained from 43 SNPs in the mentioned genes and protein variants *B, *C and *D of αS1-CN; *A1, *A2, *B, *H2 and *F of β-CN; *A and *D of αS2-CN, *A, *A1, *B, *I and *H of κ-CN and *A, *B, *C, *D, *E, *F and *H of β-LG were reconstructed. Allele and genotypic frequencies were estimated for SNPs and for protein variants; Hardy-Weinberg equilibrium and FST values were evaluated for each of the SNPs under different structuring criteria. LD values and haplotypic frequencies were estimated for caseins. The most frequent variants were CSN1S1*B (0.804), CSN2*A2 (0.509), CSN1S2*A (0.997), CSN3*A (0.679) and β-LG*B (0.657). None of the variants showed deviations from HWE, but the CSN2*A2 allele showed a slight increasing trend over time. The FST values were low (0.035) regardless of the structuring criteria. Twenty-eight CSN1S1-CSN2-CSN1S2-CSN3 haplotypes were found, 22 of them with frequencies &lt;5%; the three most frequent were BB-A1A2-AA-AA-AA-AA (16.6%), BB-A1A2-AA-AA-AA-AB (14.1%) and BB-A2A2-AA-AA-AA (10.1%). A good potential of BON cattle to produce high quality milk with functional value was reported.

Rapid detection and quantification of milk adulteration using MALDI-MS protein profiling and multivariate calibration

The adulteration of high-value milks with low-value milks is a common fraudulent practice in the dairy industry. This not only results in economic or quality prejudice, but also poses a potential threat to individuals with sensitivities or allergies. Profiling the major whey and casein milk proteins can be a crucial tool in combating this malpractice, given that each mammal species exhibits a unique protein fingerprint. In this study, we employed matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-MS) as a rapid and straightforward method for profiling proteins from bovine, caprine, and ovine milks. Subsequently, we analyzed binary mixtures of goat and sheep milks with bovine milk to establish multivariate calibration models using partial least-squares (PLS) to quantify the adulteration of goat and sheep milks. Proteins were identified according to their molecular mass (Mr) and their intensities were considered as multivariate data. To achieve satisfactory calibration and validation results, normalization through variable augmentation was necessary, enabling the quantification of bovine milk across the entire adulteration range. This uncomplicated approach based on MALDI-MS protein profiling and PLS shows significant potential for qualitative and quantitative assessments of milk adulteration.

Site specific insertion of a transgene into the murine α-casein (CSN1S1) gene results in the predictable expression of a recombinant protein in milk.

Gene loci of highly expressed genes provide ideal sites for transgene expression. Casein genes are highly expressed in mammals leading to the synthesis of substantial amounts of casein proteins in milk. The α-casein (CSN1S1) gene has assessed as a site of transgene expression in transgenic mice and a mammary gland cell line. A transgene encoding an antibody light chain gene (A1L) was inserted into the α-casein gene using sequential homologous and site-specific recombination. Expression of the inserted transgene is directed by the α-casein promoter, is responsive to lactogenic hormone activation, leads to the synthesis of a chimeric α-casein/A1L transgene mRNA, and secretion of the recombinant A1L protein into milk. Transgene expression is highly consistent in all transgenic lines, but lower than that of the α-casein gene (4%). Recombinant A1L protein accounted for 0.5% and 1.6% of total milk protein in heterozygous and homozygous transgenic mice, respectively. The absence of the α-casein protein in homozygous A1L transgenic mice leads to a reduction of total milk protein and delayed growth of the pups nursed by these mice. Overall, the data demonstrate that the insertion of a transgene into a highly expressed endogenous gene is insufficient to guarantee its abundant expression.

In vitro and in silico studies for the identification of anti-cancer and antibacterial peptides from camel milk protein hydrolysates.

Today, breast cancer and infectious diseases are very worrying that led to a widespread effort by researchers to discover natural remedies with no side effects to fight them. In the present study, we isolated camel milk protein fractions, casein and whey proteins, and hydrolyzed them using pepsin, trypsin, and both enzymes. Screening of peptides with anti-breast cancer and antibacterial activity against pathogens was performed. Peptides derived from whey protein fraction with the use of both enzymes showed very good activity against MCF-7 breast cancer with cell viability of 7.13%. The separate use of trypsin and pepsin to digest whey protein fraction yielded peptides with high antibacterial activity against S. aureus (inhibition zone of 4.17 ± 0.30 and 4.23 ± 0.32 cm, respectively) and E. coli (inhibition zone of 4.03 ± 0.15 and 4.03 ± 0.05 cm, respectively). Notably, in order to identify the effective peptides in camel milk, its protein sequences were retrieved and enzymatically digested in silico. Peptides that showed both anticancer and antibacterial properties and the highest stability in intestinal conditions were selected for the next step. Molecular interaction analysis was performed on specific receptors associated with breast cancer and/or antibacterial activity using molecular docking. The results showed that P3 (WNHIKRYF) and P5 (WSVGH) peptides had low binding energy and inhibition constant so that they specifically occupied active sites of protein targets. Our results introduced two peptide-drug candidates and new natural food additive that can be delivered to further animal and clinical trials.

The Microstructure, Rheological Characteristics, and Digestibility Properties of Binary or Ternary Mixture Systems of Gelatinized Potato Starch/Milk Protein/Soybean Oil during the In Vitro Digestion Process.

The in vitro digestibility of potato starch-based foods interacting with milk protein and soybean oil was investigated. Microstructures and rheological changes upon digestion were determined. The results showed that the addition of milk proteins (casein and whey protein) promoted gelatinized potato starch digestion, while soybean oil slowed down gelatinized potato starch digestion. A mixture of soybean oil and milk protein promoted the digestion of milk protein, while a mixture of gelatinized potato starch and milk protein inhibited the digestion of milk protein. The mixture of milk protein and/or gelatinized potato starch with soybean oil promoted the release of free fatty acids in soybean oil. The highest release rate of free fatty acids was attained by a mix of milk protein and soybean oil. The mixed samples were digested and observed with a confocal laser scanning microscope. The viscosity of the digestates was determined by a rheometer. Overall, the results demonstrated that the addition of milk protein and soybean oil had an effect on the in vitro digestibility of gelatinized potato starch and its microstructure.

Identification and evaluation of antioxidant activity of bioactive casein-derived peptides during in vitro digestion and transepithelial transport in Caco-2 cells

Dairy products are important sources of bioactive (antibacterial, antithrombotic, antihypertensive, immunomodulatory, and antioxidative) peptides, that need to be released from the target proteins. In the present study, bovine milk casein protein was hydrolyzed by pancreatin and trypsin (4 and 24 h) to evaluate the hydrolyses efficiency (ninhydrin reaction); to elucidate the sequence of casein-derived peptides (SDS-PAGE, LC-ESI-TOF-MS); and to assess the peptides antioxidant activity (DPPH and ORAC), bioactivity in silico, bioaccessibility after in vitro digestion, and bioavailablility by transepithelial transport in Caco-2 cells. The hydrolysis degree increased with the incubation time, with evident degradation of casein-derived proteins into low molecular weight peptides (<5 kDa) after 4 and 24 h incubation. Pancreatin showed higher hydrolysis degree (91%) than trypsin (21%). An increase in antioxidant activity was observed with increasing incubation time, with higher antioxidant effect observed for trypsin derived peptides (47% reduction of free radicals; 32112 μM Trolox equivalents) than pancreatin derived peptides (15% reduction; 2862 μM Trolox equivalents). A total of 69 caseins derived peptides were identified after trypsin (33 peptides) and pancreatin (36 peptides) hydrolysis for 24 h; with 26 peptides released from β-casein, 21 from α-s1-casein, 15 from α-s2-casein, and 7 from k-casein hydrolysis. Some of these peptides were described as antioxidant, anti-inflammatory, and antihypertensive. After in vitro digestion 123 peptides were found in gastric (56 peptides), duodenal (43 peptides) and colonic (24 peptides) fraction; while five peptides were bioavailable by crossing the intestinal barrier, with increased peptides’ abundance after trypsin hydrolysis treatment.

Optimizing the effect of plant protease on different properties of analog cheese containing functional corn leachate.

Cheese is produced in different flavors, textures, and forms by coagulating the milk protein casein. This study investigated the possibility of producing analog cheese by using corn steep liquor with Withania coagulans extract (WCE) and adding Eryngium planum extract (EPE) and Origanum majorana extract (OME) as functional ingredients. Different physicochemical, microbial, texture, and sensory properties of the samples were evaluated. The results obtained for moisture factor, fat, ash, water content, L*, b*, firmness, overall form, Lactobacillus and overall acceptance of the effect of all three process variables, pH, and acidity show that only the effect of WCE and OME is significant. Also, the protein of the samples was significant only on WCE and EPE (p < .001). The results showed that an increase in the levels of independent variables resulted in an increase in the amount of moisture, ash, protein, Lactobacillus, and b* and a decrease in fat, syneresis, texture properties, coliform, and lightness. Evaluation of the overall acceptance showed that consumer acceptance increased with the increase in WCE, but it initially increased and then decreased with the increase in EPE and OME levels. Finally, the samples containing 1.5% WCE, 1% EPE, and 0.5% OME were chosen as the optimized ones.

Differential behavior of Lactobacillus helveticus B1929 and ATCC 15009 on the hydrolysis and angiotensin-I-converting enzyme inhibition activity of fermented ultra-high-temperature milk and nonfat dried milk powder

Consumers' growing interest in fermented dairy foods necessitates research on a wide array of lactic acid bacterial strains to be explored and used. This study aimed to investigate the differences in the proteolytic capacity of Lactobacillus helveticus strains B1929 and ATCC 15009 on the fermentation of commercial ultra-pasteurized (UHT) skim milk and reconstituted nonfat dried milk powder (at a comparable protein concentration, 4%). The antihypertensive properties of the fermented milk, measured by angiotensin-I-converting enzyme inhibitory (ACE-I) activity, were compared. The B1929 strain lowered the pH of the milk to 4.13 ± 0.09 at 37°C after 24 h, whereas ATCC 15009 needed 48 h to drop the pH to 4.70 ± 0.18 at 37°C. Two soluble protein fractions, one (CFS1) obtained after fermentation (acidic conditions) and the other (CFS2) after the neutralization (pH 6.70) of the pellet from CFS1 separation, were analyzed for d-/l-lactic acid production, protein concentration, the degree of protein hydrolysis, and ACE-I activity. The CFS1 fractions, dominated by whey proteins, demonstrated a greater degree of protein hydrolysis (7.9%) than CFS2. On the other hand, CFS2, mainly casein proteins, showed a higher level of ACE-I activity (33.8%) than CFS1. Significant differences were also found in the d- and l-lactic acid produced by the UHT milk between the 2 strains. These results attest that milk casein proteins possessed more detectable ACE-I activity than whey fractions, even without a measurable degree of hydrolysis. Findings from this study suggest that careful consideration must be given when selecting the bacterial strain and milk substrate for fermentation.

Nearly Reversible Expansion and Shrinkage of Casein Microparticles Triggered by Extreme pH Changes.

Solvent flows in the fL/s range across the total surface of a casein microparticle cause its expansion and shrinkage. Microparticles prepared from the milk protein casein have a porous and flexible inner structure with water-filled channels and cavities. Solvent uptake occurs in two phases and results in disintegration if de-swelling is not triggered by acidification. So far, nothing is known about the reversibility of the swelling/de-swelling steps. We performed pH jump experiments between pH 11 and pH 1 on a single micro-particle and analyzed the swelling-induced size changes with system dynamics modeling. Both the swelling steps and the subsequent de-swelling process proceed reversibly and at an unchanged rate over a sequence of at least three pH exchange cycles. We observed that the duration of the first swelling step increased during the sequence, while the second step became shorter. Both of the time intervals are negatively correlated, while a statistical evaluation of only one swelling cycle for an ensemble of microparticles with different stabilities did not reveal any significant correlation between the two parameters. Our results indicate that the pH-induced swelling/shrinkage of casein microparticles is, to a large extent, reversible and only slightly influenced by the acid-induced decomposition of colloidal calcium phosphate.

Soft Matter Engineering for the Fabrication of Functionalized Casein Microparticles

AbstractThe structural features of the most abundant milk protein casein can be exploited for the targeted production of microparticles with desired properties. After the addition of pectin, the so‐called casein micelles reversibly form aggregates of several µm in size, which change their shape due to shear and elongation forces. The soft properties are crucial for the subsequent solidification into stable casein microparticles during film drying. Based on pH‐dependent experiments, we show how we analyze and tailor the swelling and disintegration properties of the microparticles for future controlled and instant release applications.

Review: Genetic and protein variants of milk caseins in goats.

The milk casein genes in goats, are highly polymorphic genes with numerous synonymous and non-synonymous mutations. So far, 20 protein variants have been reported in goats for alpha-S1-casein, eight for beta-casein, 14 for alpha-S2-casein, and 24 for kappa-casein. This review provides a comprehensive overview on identified milk casein protein variants in goat and non-coding DNA sequence variants with some affecting the expression of the casein genes. The high frequency of some casein protein variants in different goat breeds and geographical regions might reflect specific breeding goals with respect to milk processing characteristics, properties for human nutrition and health, or adaptation to the environment. Because protein names, alongside the discovery of protein variants, go through a historical process, we linked old protein names with new ones that reveal more genetic variability. The haplotypes across the cluster of the four genetically linked casein genes are recommended as a valuable genetic tool for discrimination between breeds, managing genetic diversity within and between goat populations, and breeding strategies. The enormous variation in the casein proteins and genes is crucial for producing milk and dairy products with different properties for human health and nutrition, and for genetic improvement depending on local breeding goals.

Effects of the maize-derived protein zein, and the milk proteins casein, whey, and α-lactalbumin, on subjective measures of satiety and food intake in normal-weight young men

Protein is considered to be the most satiating food macronutrient and the satiating effect may be dependent on the source of the protein. The maize-derived protein zein and milk protein casein have been shown previously to lower stomach emptying rate more than dairy whey protein, but the effect of zein on satiety has not been evaluated. The objective was to compare the satiating effects of zein and casein, with whey protein and its protein component α-lactalbumin. The study was a randomised crossover design with thirteen normal-weight men (mean age 27.8 years and mean BMI 24.4 kg/m2) consuming isoenergetic (∼4000 kJ, ∼990 kcal) preload mixed meals enriched with Zein, Casein, whey protein isolate (Whey), α-lactalbumin (ALac), or maltodextrin carbohydrate (Carb). Consumption of an ad libitum standardised test meal of chicken fried rice and water provided 360 min following ingestion of the preload meal was measured, and subjective feelings of appetite (hunger, fullness, desire to eat, and prospective food consumption) were assessed using 100-mm visual analogue scales (VAS). There were no differences among the five preload mixed meals in the amount of chicken fried rice consumed at the ad libitum test meal (mean ± sem: 531.6 ± 35.0 g, p = 0.47) or total (preload + test meal) energy intakes (mean ± sem: 5780.5 ± 146.0 kJ, p = 0.29). The subjective VAS appetite ratings and total area under the curve responses for hunger, fullness, desire to eat, and prospective food consumption, were not different following consumption of all five preload mixed meals (p > 0.05). The findings indicate that the effects of zein and casein on satiety were not different from the satiating effects of whey protein and α-lactalbumin.

Determination of Anti Tumors Level of Caseın in Human Adenokarsinom Cell

Introduction: In this study, it was aimed to investigate the cytotoxicity and antitumoral activity of casein proteins obtained from bovine milk on colorectal cancer cells (Caco-2). Methods: Milk were fractionated as casein proteins. Antitumor activity levels of different concentrations of lyophilized proteins were measured by MTT viability test on cells and IC50 values were determined. Conclusion: Serial dilutions of casein proteins in milk, starting with 3.200 µg/ml and decreasing, were incubated with Caco-2 cells for 24, 48 and 72 hours. When incubated with bovine casein Caco-2 cells, cell viability: IC50 value found in the 24 hour run 9.172 µg/ml 69.21% at 3.200 µg/ml; IC50 at 48 hours was 8.927 cell viability; 58.5%; The IC50 value at 72 hours was 8.043 µg/ml, and cell viability was found to be 51.91%. It was concluded that bovine casein proteins have limited antiproliferative and anticarcinogenic effects on Caco-2 cancer cell lines.

Influence of milk proteins on the adhesion and formation of Bacillus sporothermodurans biofilms: Implications for dairy industrial processing

Bacillus sporothermodurans is a producer of highly heat-resistant spores, which is a problem for the dairy industry worldwide. In this work, we studied the attachment and biofilm formation on stainless steel surfaces in contact with milk proteins (casein and whey). The results of the biofilm formation were obtained by the plate count method. In conjunction, biofilms (niches and cell sizes) were measured by scanning electron microscopy (SEM) on the 1 st , 5 th and 10 th day. X-ray photoelectron spectroscopy (XPS) was a complementary technique that probed the conditioning of the substrate surface after 24 h. The milk proteins affected the total bacterial count in the samples, and also influenced the biofilm architecture. Sessile cell counts varied from 4.3 log CFU/cm 2 in the attachment stage to up to 10.2 log CFU/cm 2 in the matured biofilm stage. The spore counts varied from <1.4 ± 0.02 (1 st day) to 3.7 log ± 1.1 log spores/cm 2 (10 th day). The control group at the biofilm maturation stage (5 th day) had higher niches than at the cell fixation (1 st day) and was different from the other groups that showed active dispersion in the presence of milk proteins. The same occurred on the 10 th day for the control group. Circular patterns in the biofilm with casein, passive dispersal and elongated cells were also observed. Enzyme treatment and disinfectants may be used to remove or reduce biofilm formation. • This is the first report on the B. sporothermodurans biofilm life cycle. • The distributions of B. sporothermodurans biofilms were influenced by milk proteins. • Active and passive dispersal were observed in the biofilm life cycle. • Enzymatic and chemical treatments are efficient for the removal of biofilm. • Eventual pitting or corrosion and biofilm formation at the air-liquid interface were observed.

Inorganic food additive nanomaterials alter the allergenicity of milk proteins

While inorganic nanomaterials are copiously incorporated in food products, their impact on the allergenicity food proteins is largely unknown. This study analyzed the effect of widely used food additive nanomaterials (silica and titania) on the antigenicity and allergenicity of milk proteins (β-lactoglobulin and casein) and skimmed milk. Changes in the antigenicity of milk proteins in the presence of dietary nanomaterials were identified using an indirect-ELISA assay, while the change in allergenicity was studied using mast cell (LAD2) sensitized using allergic human sera. Results showed an enhancement in the allergenicity of milk proteins/skimmed milk interacted with particles (both silica and titania). Similarly, mast cell degranulation (a proxy for allergenicity) was higher when exposed to particle interacted skim milk where nanomaterials of titania showed the highest effect, and this tendency was retained even after subjecting to simulated gut digestion. Particles induced alterations in the structure of milk proteins, as evidenced by our studies, are reasoned to expose epitopes that increase allergenicity of milk proteins.

Casein Oligochitosan-Glycation by Transglutaminase Enhances the Anti-Inflammatory Potential of Casein Hydrolysates to the Lipopolysaccharide-Stimulated IEC-6 Cells.

In this study, milk protein casein was glycated by oligochitosan through the catalysis of transglutaminase (TGase) and then hydrolyzed by trypsin. The obtained glycated casein hydrolysates (GCNH) were assessed for their anti-inflammatory activities, using the lipopolysaccharide (LPS)-stimulated rat intestinal epithelial cells (IEC-6) as cell models and the casein hydrolysates (CNH) without TGase catalysis as controls. The results showed that GCNH had oligochitosan incorporation and thus possessed a glucosamine content of 5.74 g/kg protein. In general, GCNH at dose levels of 25–100 μg/mL could elevate IEC-6 cell growth, and at dose levels of 25–50 μg/mL, they were also able to alleviate the LPS-induced cytotoxicity by increasing cell viability efficiently. Although LPS caused clear inflammation in the LPS-stimulated cells, GCNH were capable of reducing the secretion of three pro-inflammatory mediators including interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α, or promoting the secretion of two anti-inflammatory mediators like IL-10 and transforming growth factor-β, demonstrating their anti-inflammatory activities to the stimulated cells. Moreover, GCNH also could down-regulate the expression of three inflammation-related proteins including TLR4, p-p38, and p-p65 in the stimulated cells, and thus possessed a capacity to suppress the phosphorylation of p38 and p65 proteins as well as to inactivate the NF-κB and MAPK signaling pathways. Additionally, a higher GCNH dose level consistently led to higher anti-inflammatory effect in the cells, while GCNH were always more potent than CNH at performing anti-inflammatory function targets. It is thus suggested that the TGase-catalyzed casein oligochitosan-glycation could enhance the anti-inflammatory activities of casein hydrolysates efficiently. TGase-catalyzed protein glycation thus might enhance the healthcare function of protein ingredients in the body.