Development of livestock feed additives using porcine blood cells.

Livestock blood discarded during slaughtering has potentially valuable components such as plasma proteins and haemoglobin. Plasma is used as a feed additive following processing via different methods, including spray drying, whereas blood cells have been underutilized. In this study, we developed haemoglobin hydrolysate (HH) and iron-enriched residue (IER) from porcine blood cells and investigated whether their oral administration regulates the immune system and gut microbiota in growing rats. Twenty-one Sprague-Dawley male rats (n = 7) were used during a 4-week trial and were fed a control, HH or IER diet. The ratio of beneficial bacteria such as Lactobacillus and Akkermansia strains increased in rats fed HH or IER diets. Moreover, compared with the control group, the IER group had an elevated ratio of Lactobacillus to Enterobacteria, which is regarded as an index of beneficial aspect in the gut. Phagocytosis of peripheral blood leucocytes was higher in the HH and IER groups than in the control group. The level of plasma immunoglobulin G increased to approximately 72.7 mg/ml and 152.0 mg/ml in the HH and IER groups, respectively, which was significantly (p < 0.05) higher than that in the control group. These results confirm that HH and IER developed in this study may be a potential additive for animal feeds.

Correlation of attention deficit hyperactivity disorder with gut microbiota according to the dietary intake of Korean elementary school students.

We investigated the impact of dietary patterns on the gut microbiota and concentration of short-chain fatty acids in the feces of Korean elementary school students. The dietary intake and ADHD assessment of 40 Korean elementary school students were analyzed using a dish-based semi-quantitative food frequency questionnaire. Analysis of gut microbiota and short-chain fatty acids composition were performed using the real-time polymerase chain reaction, metagenomics, and gas chromatography methods. The dietary patterns of participants were divided into four groups: healthy, processed food, fish and shellfish, and meat. The participants were also divided into two groups according to their ADHD scores: 0-30, control group; over 30, ADHD group. The ADHD score of the processed food group was significantly higher than that of the healthy group. The processed food and ADHD groups showed significantly higher abundance of harmful bacteria, such as the Enterobacter, Escherichia coli, and Clostridium strains, and markedly lower abundance of beneficial bacteria, such as the Bifidobacterium and Ruminococcus strains, than the control group. The heat maps of metagenomics indicated that each group was separated into distinct clusters, and the processed food and ADHD groups showed significantly lower α-diversity of gut microbiota than the control group. In these groups, the concentration of acetate or butyrate in the feces was significantly lower than that in the control group. These results may indicate that imbalanced diets can disturb the colonic microbial balance and are likely to become a potential risk factor for the prevalence of ADHD.

Butyrate modulates mucin secretion and bacterial adherence in LoVo cells via MAPK signaling.

Short-chain fatty acids contribute to normal bowel function and prevent bacterial infections. In particular, butyrate is a promising candidate that plays an important role in regulating the functional integrity of the gastrointestinal tract by stimulating mucin secretion. We investigated whether butyrate treatment modulates mucin secretion and bacterial adherence in LoVo cells. In addition, the possible signaling pathways were also examined in connection with the upregulation of mucin secretion. The results showed that butyrate induced mucin secretion in LoVo cells, resulting in the inhibition of Escherichia coli adhesion by increasing the adherence of Lactobacillus acidophilus and Bifidobacterium longum. The gene expression analysis suggests that mitogen-activated protein kinase (MAPK) signaling pathways including Cdc42-PAK pathway appears to be involved in stimulating mucin secretion. More importantly, butyrate induced the increased actin expression and polymerization in LoVo cells, which could be attributable to the Cdc42-PAK signaling pathway, implicated in actin cytoskeleton and mucin secretion. Our results provide a molecular basis in modulating bacterial adherence and the MAPK signaling pathway for the improved homeostasis of colonic epithelial cells.

Imbalanced dietary intake alters the colonic microbial profile in growing rats.

An imbalanced dietary intake is associated with alteration of intestinal ecosystem. We investigated the impact of imbalanced diets on colonic microbiota, concentrations of short chain fatty acid in colonic digesta and serum immunoglobulins (Igs) of growing rats. Compared to the control diet, consuming diets high in fat, sucrose, or processed meat, or low in iron, increased the abundance of the pathogenic bacteria such as Clostridium, Escherichia coli, and Salmonella species, and decreased the beneficial bacteria, like Bifidobacteria, Lactobacillus, Akkermansia, Phascolarctobacterium, Alistipes, and butyrate producing species of bacteria in the colon of growing rats. The heatmap of metagenomics indicated that each group was separated into distinct clusters, and the ID group in particular, showed significantly (P < 0.01) reduced alpha diversity of colonic microbiota in comparison to the control group. All experimental groups showed significantly (P < 0.05 or P < 0.01) decreased concentration of acetate and butyrate in the colonic digesta and lower levels of serum IgG or IgA, compared to the control. These results indicated that the imbalanced dietary intake negatively altered intestinal ecosystem and immunity.

Enzymatic Hydrolysis of Egg White Protein Exerts a Hypotensive Effect in Spontaneously Hypertensive Rats.

This study was conducted to investigate the hypotensive effect of egg white protein (EWP) hydrolysate (EWH) in spontaneously hypertensive rats (SHRs). The hydrolysis of EWP was effectively performed with a combination of 0.5% bromelain and 1% papain at 50°C for 60 min. The resulting hydrolysate did not elicit an allergic reaction as confirmed by human mast cell activation test. The systolic and diastolic blood pressures of the SHRs fed the EWH diet were observed to be significantly or numerically lower than those of the other groups during the experimental period of 28 d. EWH treatment significantly (p<0.05) upregulated the nitric oxide levels in hCMEC/D3 cells and the plasma of the SHRs compared to those in the control. Moreover, EWH ingestion significantly (p<0.01) reduced the plasma angiotensin II level of the SHRs compared with that in the control. In conclusion, beyond its basic nutritional value, EWH prevents and manages hypertension, and thus can be an invaluable resource for functional food development.

Evaluation of the Digestibility of Korean Hanwoo Beef Cuts Using the in vitro Physicochemical Upper Gastrointestinal System.

The aim of this study was to investigate the digestibility of different Korean Hanwoo beef cuts using an in vitro digestion model, in vitro physicochemical upper gastrointestinal system (IPUGS). The four most commonly consumed cuts - tenderloin, sirloin, brisket and flank, and bottom round - were chosen for this study. Beef samples (75 g) were cooked and ingested into IPUGS, which was composed of mouth, esophagus, and stomach, thereby simulating the digestion conditions of humans. Digested samples were collected every 15 min for 4 h of simulation and their pH monitored. Samples were visualized under a scanning electron microscope (SEM) to examine changes in the smoothness of the surface after digestion. Analysis of the amino acid composition and molecular weight (MW) of peptides was performed using reverse-phase high performance liquid chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis, respectively. Following proteolysis by the gastric pepsin, beef proteins were digested into peptides. The amount of peptides with higher MW decreased over the course of digestion. SEM results revealed that the surface of the digested samples became visibly smoother. Total indispensable and dispensable amino acids were the highest for the bottom round cut prior to digestion simulation. However, the total amount of indispensable amino acids were maximum for the tenderloin cut after digestion. These results may provide guidelines for the elderly population to choose easily digestible meat cuts and products to improve their nutritional and health status.

Purification and Anti-pathogenic Properties of Immunoglobulin Concentrates from Porcine Blood.

During slaughtering, animal blood is typically discarded, resulting in water pollution. However, this discarded blood has valuable components, such as immunoglobulin (Ig). Although several studies have been conducted to develop methods for effective recycling of slaughterhouse blood, they have not been commercially utilized in Korea. Here, we extracted an Ig-rich fraction from porcine blood that was then subjected to various in vitro tests, including pathogen growth inhibition, antigenic cross-reactivity, and anti-toxin activity. The porcine immunoglobulin concentrate (PIC) was effectively purified by eliminating other components, such as albumin, and consisted of approximately 63.2±2.9% IgG and 7.2±0.4% IgM on a protein basis. The results showed that it significantly suppressed the growth of pathogenic bacteria, and bound to all tested pathogens, including both gram-positive and gram-negative species, although the degree of activity differed according to strain. The PIC bound to two types of lipopolysaccharide (LPS) obtained from Escherichia coli O111:B4 and Salmonella enterica serotype typhimurium in a concentration-dependent manner. In addition, the PIC restored the proliferation activity of the lymphoblast K-562 cells when co-incubated with pathogenic LPS. These results confirm that the PIC prepared in this study is a potentially valuable functional food material or diet supplement as an alternative to antibiotics that can protect animals from pathogenic bacteria.

Hypoallergenic and Physicochemical Properties of the A2 ß-Casein Fractionof Goat Milk.

Goat milk has a protein composition similar to that of breast milk and contains abundant nutrients, but its use in functional foods is rather limited in comparison to milk from other sources. The aim of this study was to prepare a goat A2 ß-casein fraction with improved digestibility and hypoallergenic properties. We investigated the optimal conditions for the separation of A2 ß-casein fraction from goat milk by pH adjustment to pH 4.4 and treating the casein suspension with calcium chloride (0.05 M for 1 h at 25°C). Selective reduction of ß-lactoglobulin and αs-casein was confirmed using sodium dodecyl sulphate-polyacrylamide gel electrophoresis and reverse-phase high-performance liquid chromatography. The hypoallergenic property of A2 ß-casein fraction was examined by measuring the release of histamine and tumor necrosis factor alpha from HMC-1 human mast cells exposed to different proteins, including A2 ß-casein fraction. There was no significant difference in levels of both indicators between A2 ß-casein treatment and the control (no protein treatment). The A2 ß-casein fraction is abundant in essential amino acids, especially, branched-chain amino acids (leucine, valine, and isoleucine). The physicochemical properties of A2 ß-casein fraction, including protein solubility and viscosity, are similar to those of bovine whole casein which is widely used as a protein source in various foods. Therefore, the goat A2 ß-casein fraction may be useful as a food material with good digestibility and hypoallergenic properties for infants, the elderly, and people with metabolic disorders.

Hydrolysis by Alcalase Improves Hypoallergenic Properties of Goat Milk Protein.

Goat milk is highly nutritious and is consumed in many countries, but the development of functional foods from goat milk has been slow compared to that for other types of milk. The aim of this study was to develop a goat milk protein hydrolysate (GMPH) with enhanced digestibility and better hypoallergenic properties in comparison with other protein sources such as ovalbumin and soy protein. Goat milk protein was digested with four commercial food-grade proteases (separately) under various conditions to achieve the best hydrolysis of αs -casein and ß-lactoglobulin. It was shown that treatment with alcalase (0.4%, 60℃ for 30 min) effectively degraded these two proteins, as determined by SDS-PAGE, measurement of nonprotein nitrogen content, and reverse-phase high-performance liquid chromatography. Hydrolysis with alcalase resulted in a significant decrease in ß-lactoglobulin concentration (almost to nil) and a ~40% reduction in the level of αs-casein. Quantification of histamine and TNF-α released from HMC-1 cells (human mast cell line) showed that the GMPH did not induce an allergic response when compared to the control. Hence, the GMPH may be useful for development of novel foods for infants, the elderly, and convalescent patients, to replace cow milk.

In Vitro Effects of Dietary Inulin on Human Fecal Microbiota and Butyrate Production.

Administration of dietary fibers has various health benefits, mainly by increasing numbers of beneficial bacteria and enhancing production of short-chain fatty acids in the colon. There has been growing interest in the addition of dietary fiber to human diet, due to its prebiotic effects. This study aimed to evaluate the prebiotic activity of inulin using an in vitro batch fermentation system with human fecal microbiota. Fermentation of inulin resulted in a significantly greater ratio of Lactobacillus or Bifidobacteria to Enterobacteria strains as an index of healthy human intestine and elevated butyrate concentration, which are related to improvement of gut health.

Butyrate modulates bacterial adherence on LS174T human colorectal cells by stimulating mucin secretion and MAPK signaling pathway.

BACKGROUND/OBJECTIVES: Fermentation of dietary fiber results in production of various short chain fatty acids in the colon. In particular, butyrate is reported to regulate the physical and functional integrity of the normal colonic mucosa by altering mucin gene expression or the number of goblet cells. The objective of this study was to investigate whether butyrate modulates mucin secretion in LS174T human colorectal cells, thereby influencing the adhesion of probiotics such as Lactobacillus and Bifidobacterium strains and subsequently inhibiting pathogenic bacteria such as E. coli. In addition, possible signaling pathways involved in mucin gene regulation induced by butyrate treatment were also investigated. MATERIALS/METHODS: Mucin protein content assay and periodic acid-Schiff (PAS) staining were performed in LS174T cells treated with butyrate at various concentrations. Effects of butyrate on the ability of probiotics to adhere to LS174T cells and their competition with E. coli strains were examined. Real time polymerase chain reaction for mucin gene expression and Taqman array 96-well fast plate-based pathway analysis were performed on butyrate-treated LS174T cells. RESULTS: Treatment with butyrate resulted in a dose-dependent increase in mucin protein contents in LS174T cells with peak effects at 6 or 9 mM, which was further confirmed by PAS staining. Increase in mucin protein contents resulted in elevated adherence of probiotics, which subsequently reduced the adherent ability of E. coli. Treatment with butyrate also increased transcriptional levels of MUC3, MUC4, and MUC12, which was accompanied by higher gene expressions of signaling kinases and transcription factors involved in mitogen-activated protein kinase (MAPK) signaling pathways. CONCLUSIONS: Based on our results, butyrate is an effective regulator of modulation of mucin protein production at the transcriptional and translational levels, resulting in changes in the adherence of gut microflora. Butyrate potentially stimulates the MAPK signaling pathway in intestinal cells, which is positively correlated with gut defense.

One-step production of superhydrophobic coatings on flat substrates via atmospheric Rf plasma process using non-fluorinated hydrocarbons.

This paper describes the direct deposition of hydrocarbon coatings with a static water contact angle higher than 150 using simple C6 hydrocarbons as a reactive gas in helium plasma generated in ambient air without any preroughening of the silicon (100) substrate. The film morphology and hydrophobicity are found to strongly depend on the structure of the reagent hydrocarbon. The films deposited with n-hexane and cyclohexane exhibited relatively smooth morphology and the water contact angle was only ∼95°, similar to polypropylene. When benzene was used as a main reactive gas, the deposited film surface showed nanoscale textured morphology and superhydrophobicity with a water contact angle as high as 167°. Because the plasma is generated in air, all films show some degree of oxygen incorporation. These results imply that the incorporation of a small amount of oxygenated species in hydrocarbon films due to excitation of ambient air is not detrimental for superhydrophobicity, which allows the atmospheric rf plasma with the benzene precursor to produce rough surface topography needed for superhydrophobicity.