Pepsin Activity Research Articles

Effects of dietary fish meal replacement by Periplaneta americana meal on growth, metabolism, intestinal health and resistance against Aeromonas hydrophila of Micropterus salmoides

An 8-week culture experiment was conducted to investigate the effects of partial replacement of fish meal (FM) with Periplaneta americana meal (PAM) on growth performance, body composition, serum biochemical parameters, intestinal health and resistance against Aeromonas hydrophila of largemouth bass, Micropterus salmoides. A total of 450 healthy largemouth bass with an initial body weight (IBW) of 14.55±0.09 g were randomly divided into five groups. Five experimental diets were prepared by replacing 0 % (P0), 15 % (P15), 30 % (P30), 45 % (P45) and 60 % (P60) of the FM in the basal diet with PAM, respectively. The results showed that final body weight (FBW), weight gain rate (WGR), and specific growth rate (SGR) were no significant difference in P15 and P30 compared with the control group (P > 0.05). Crude protein of whole fish body was significantly increased and crude lipid was significantly decreased in all PAM groups (P < 0.05). In the P30, the antioxidant enzyme activities were significantly higher than those in the control and other experimental groups, and malondialdehyde (MDA) was significantly decreased (P < 0.05). The activities of α-amylase and pepsin increased significantly in P15 and P30 and decreased significantly in P45 and P60 (P < 0.05). Lipase was significantly increased in all groups except P15 than in the control group (P < 0.05). There were no significant differences in villus height (VH), crypt depth (CD), and muscle thickness (MT) among all groups (P > 0.05), but villus width (VW) was significantly decreased in P45 and P60 (P < 0.05). The relative expression of IL-10 and TGF-β1 were first up-regulated and then down-regulated, while IL-8 was opposite (P < 0.05). The relative expression of Casp3, Casp9 and IFN-γ were significantly up-regulated in P45 and P60 (P < 0.05). After injecting A. hydrophila, all groups of largemouth bass experienced mortality, but cumulative mortality was lowest in the P30. In conclusion, PAM can partially replace FM in the diet of juvenile largemouth bass. The replacement of no more than 30 % is unlikely to adversely affect growth performance, and can increase the crude protein content of the whole fish body, and is beneficial to intestinal health.

Influence of lecithin on the digestibility and immunoreactivity of β-conglycinin

A growing body of evidence suggests that protein structure can be uniquely altered by lipids. However, the role of lecithin on the digestibility and immunoreactivity of allergenic protein remains poorly understood. Therefore, we investigated the impact of different protein/lecithin (P:L) ratios (5:1, 10:1, 20:1, 30:1 and 40:1) on the digestion properties, peptide profiles and immunoreactivity of β-conglycinin in vitro simulated gastrointestinal digestion. Results revealed that lecithin addition increased the degree of hydrolysis and digestion rate of β-conglycinin during digestion in a dose-dependent manner, which may be attributed to the enhancement of pepsin activity. Additionally, the addition of lecithin contributed to the dispersion and homogeneity of β-conglycinin digestion products. When the ratio of lecithin to β-conglycinin was 1:10, the obtained β-conglycinin digestion products exhibited the highest DH value (23.80 %) and the lowest antigenicity (52.69%) in the intestinal digestion. Peptidomics and immunoinformatic analysis further confirmed that PL-10 disrupted 18/30, 15/44, and 37/75 epitopes in the α, α′, and β subunits of β-conglycinin, respectively. These results suggested that lecithin changed the cleavage preferences of β-conglycinin, and promoted the disruption of allergic epitopes, specifically for the β subunit. The study can contribute to our understanding of the role of lecithin in the digestion properties and immunoreactivity of allergenic protein.

Effects of dietary rosemary (Rosmarinus officinalis) leaf powder and Bacillus subtilis on the growth performance, digestive enzymes, antioxidant and immune-related gene expression, and resistance to Aeromonas hydriphila in common carp, Cyprinus carpio

Abstract Due to increasing demand about the application of natural feed additives in aquaculture, the efficacy of separate and the combination of rosemary (Rosmarinus officinalis) and B. subtilis on the growth performance, blood indices, innate immunity, antioxidant defense, digestive enzymes activity, the expression levels of genes with immunity modulation, as well as the act of resisting against Aeromonas hydriphila, in common carp, C. carpio is reported. Fish (44.63 ± 0.04 g) were divided into four treatments including the T0 (control treatment) fed with basal diet, T1 fed with rosemary at the rate of 3 %, T2 fed with 1.5×107 CFU g-1 B. subtilis, and T3 fed with a combination of rosemary at the rate of 3 % and 1.5×107 CFU g-1 B. subtilis. After 56 days enriched feed with B. subtilis and combination of rosemary and B. subtilis caused significant improvement in weight gain (WG %), feed conversion ratio (FCR), hematological and biochemical indices, the innate immune responses including in total Ig level, lysozyme and ACH50 activities as well as the chymotrypsin, pepsin, protease, and amylase activities correlate to the control group (P&lt;0.05). Additionally, the serum CAT and GPX activity were significantly enhanced in the T3 group (P&lt;0.05). The mRNA expression of CAT and LYS genes significantly increased in T2 and T3 contrary to T0 (p&lt;0.05). Fish challenged with A. hydrophila had variable survival and the best survival was in the T3. In summary, combination the rosemary and B. subtilis enhanced immune parameters and the ability of fish to cope with A. hydrophila as a fish pathogen. These findings indicated the potential of these substances as a promising feed additive.

The impact of host genetics on microbe assemblages in three types of mrigal carp (Cirrhinus mrigala)

Host–microbiota interactions are molecular and physical interactions between the microbiota and the host that play significant roles in the lifespan of animals. In the present study, two types of gynogenetic mrigal carp (GMCC and GMCW) and the original maternal mrigal carp (MC) were used as a models to investigate host–microbiota interactions. The two types of gynogenetic mrigal carps showed faster growth and better cold tolerance than the MC fish. Compared to those in MC, a large number of goblet cells and higher activity of cellulase and pepsin were detected in the intestine of gynogenetic fish. The composition and abundance of the microbial communities significantly differed among the three types of mrigal carp and water, with Proteobacteria, Firmicutes and Chloroflexi being the most dominant microbes in the GMCC, GMCW and MC fish, respectively. Pseudomonas, Lactococcus and Defluviicoccus were the biomarker genera in the GMCC, GMCW and MC fish. Network analysis revealed no relationships between fish and water, but the dominant genera was positively correlated with the abundance of certain enzymes. Functional analysis revealed the dominant genera associated with amino acid metabolism, fatty acid metabolism and bile acid biosynthesis. Our results suggested that host genetics may affected gut microbe assembly and that the specific metabolic functions of gut microbes may contribute to growth performance through the microbe-gut-liver axis.

Influence of physicochemical factors on the specific activity of proteases used in cheesemaking biotechnology

The formation of a clot under the influence of proteolytic enzymes is an essential stage in cheesemaking technology. Depending on their composition, milk-clotting enzyme preparations have a different effect on the quality characteristics of the clot, whey separation, and yield of the finished product. At present, the Russian market offers a wide range of domestic and imported milk-clotting enzyme preparations. In this connection, the choice of a milk coagulant by cheese makers should be carried out in accordance with the cheese group produced by the enterprise. The milk-clotting activity of enzyme preparations depends not only on the active acidity of the mixture, but also on the temperature of the solution used to obtain them. It is the solution temperature that has a determining effect on the duration of the coagulation process and, therefore, the quality of the formed clot. Research into the specific activity of milk-clotting enzyme preparations, obtained in a working solution with different active acidity (pH from 4.0 to 8.0), showed their milk-clotting activity to decrease significantly upon pH shifting to the alkaline region. Application of solutions with a pH of 4.0 led to an increase in the milk-clotting activity of bovine pepsin, VNIIMS SG-50 rennet-bovine preparation, and rennet enzyme by 94, 72, and 36%, respectively, relative to the control. Bovine pepsin demonstrated the highest sensitivity to fluctuations in active acidity. A study into the effect of substrate temperature in the range from 30 to 40 °C established rennet to be the most sensitive preparation to temperature changes. It is concluded that a properly selected enzyme composition is an effective means for affecting the quality of cheese products.

Quantitatively characterize the response of the hybrid grouper (Epinephelus fuscoguttatus ♀ × Epinephelus lanceolatus ♂) under elevated temperature stress

Global warming may affect the health of marine species. However, the collected information on quantitative assessment of response in fish under elevated temperature is poorly defined. The present study aimed to quantitatively evaluate the effects of the hybrid grouper (Epinephelus fuscoguttatus ♀ × Epinephelus lanceolatus ♂) under elevated temperature (33 °C and 36 °C, ET1 and ET2) stress for 14 days. As endpoints, we examined changes in body growth, hemato-immunological parameters, liver oxidative stress markers, as well as changes of the stomach digestive enzymes. Compared to the control, the body weight was significantly decreased in ET2 group for 14 d exposure, and a remarkable change of differential leukocyte counts of the fish was observed in ET1 group at 3 d and in ET2 group at 14 d. The respiratory burst activity of the hybrid grouper leukocytes markedly decreased in the treatment groups after 14-d exposure. Overall, the antioxidant enzyme activities and transcriptional levels of superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), and glutathione peroxidase (GSH-PX) were markedly inhibited in the liver for 3-d and 14-d exposure. The expression levels of nf-κb mRNA were significantly inhibited while the expression levels of atp2b1 mRNA were significantly induced after 14-d exposure. The activities of pepsin and lipase in the stomach were significantly reduced. In addition, an innovative hazard classification system (ET-HCS) was developed to quantitatively characterize the stress response of the fish following elevated temperature treatments. The stress level of ET2 group for 14-d exposure was ranked as level IV (high stress), and the other treatments were ranked as level II (low stress). Taken together, the findings of this study further extend our understanding of quantitative assessment of response in fish under high-temperature stress, which provides valuable information for improving countermeasures of mariculture industry.

491 Processing and stomach pH influenced in vitro gastric protein solubilization kinetics in sunflower meal

Abstract The study aimed to evaluate the effect of processing and pH on gastric protein solubilization kinetics in sunflower meal (SFM) products using porcine digestion simulation model. The SFM samples were derived from diverse processing approaches: 1) dehulled-cold pressed (SFM_CP), 2) dehulled-solvent-extracted (SFM_SE), and 3) application of electrostatic separation technology on SFM_SE to generate high crude protein (CP) version (SFM_SEP). Soybean meal (SBM) was included for reference comparison (Table 1). Considering the complex factors influencing retention time, gastric pH, and pepsin secretion, the study employed multiple sampling times, and pH levels. A total of 500 mg CP equivalent of each sample (in triplicates) was mixed with 25 mL of 0.1 M phosphate buffer (pH 6.0) and 2 mL of pepsin (27 mg/mL). The pH was adjusted to either 1.5, 2.5, 3.5 or 4.5 using 1 M HCl and placed in incubator shaker (200 rpm) at 39°C. Samples were taken at 0, 30, 60, and 90 min, snap-frozen in liquid nitrogen, stored frozen, later thawed at 4°C, centrifuged (15,000 × g for 10 min) and supernatants analyzed for CP. The data were analyzed using Proc GLIMIXX in SAS while Gompertz equation was used to describe the gastric protein solubilization kinetics. The model included the fixed effects of protein source or processing, stomach pH and their interaction on the gastric CP solubility at 90 min and gastric protein solubility kinetics (A-maximum solubilized CP, Wo- initial CP solubility, Ku- solubilization rate, Ti- time to inflection). The analyzed CP SBM, SFM_CP, SFM_SE, and SFM_SEP, were 514.4, 524.4, 393.5, and 484.2 g/kg DM, respectively. Gastric CP solubility increased with lower pH and longer incubation time (P &amp;lt; 0.05), which corresponded well with the optimum pH of pepsin activity around 2. Compared with SFM, SBM had the least (P &amp;lt; 0.05) gastric CP solubility at pH 4.5 and 3.5, and the least predicted maximum gastric CP solubility (A) at pH 4.5, 3.5 and 2.5 (P &amp;lt; 0.05; Table 2). SFM_CP showed the greatest predicted initial gastric CP solubility at 0 h (Wo) and A (P &amp;lt; 0.05). Electrostatic fractionation of SFM_SE (SFM_SEP) enhanced gastric CP solubility at pH 3.5 and 2.5 and reduced the solubility time to inflection point (Ti; P &amp;lt; 0.05). In conclusion, data indicated that cold pressing SFM increased gastric CP solubility relative to solvent extraction. However, electrostatic fractionation of SFM_SE improved protein gastric solubility. While SBM remains a standard, incorporating the studied sunflower meals with their fast CP digestion kinetics in feed formulation could improve protein utilization in young animals with high stomach pH. In vivo studies, particularly on ileal amino acid digestibility, are crucial to validate these findings. Sunflower meal tables_FMN.pdf

Effects of water temperature on growth, health and digestive processes of the thicklip grey mullet Chelon labrosus

This study aimed to identify the effects of water temperature on the rearing of the omnivorous teleost Chelon labrosus and to determine the optimal temperature for the culture of this interesting candidate for the diversification of European aquaculture. The fish (initial weight 26.81 ± 6.2 g) were held at four water temperatures (18, 22, 26 and 30 °C) in triplicate for 90 days in 100 L recirculating tanks (n = 14 / tank). The effects of said temperatures were assessed on growth, intestinal health and the digestive processes of the fish. At 18 °C, energy consumption was low, leading to a high accumulation of reserves and slow growth, a typical overwintering strategy. The best growth results were obtained at 22 °C, followed closely by 26 °C, which resulted in low energy reserves. The fact that the fish at 26 °C grew less than the ones at 22 °C while having similar energy consumption, suggested that this temperature was close to the end of the optimal range, but no signs of stress were detected. However, at 30 °C the fish had lower energy reserves but grew significantly less, which can be considered indicative of stress. Health impairment at this temperature was confirmed by the epithelial lesions found in this group. The growth and energy availability of the different groups is discussed in the light of the activities of α-amylase, pepsin and alkaline proteases. As an integration of all these results, a quadratic regression model resulting from growth values (SGR) in relation to water temperature was performed, and it allowed predicting an actual optimal water temperature of 22.8 °C for growth of C. labrosus. This is the first assessment of the optimal water temperature for the culture of C. labrosus, and it will be highly valuable for the development of its intensive culture.

Conformational alterations and functional changes of pepsin induced by a novel food supplement tetrahydrocurcumin: Multispectral techniques and computer simulations

Tetrahydrocurcumin (THC), as a novel food supplement, has generated significant interests for its potential impact on health and nutrition. Pepsin serves as the primary enzyme involved in the digestive mechanism. This research investigated the conformational and functional alterations of pepsin induced by THC using multispectral techniques and computer simulations. The results showed that THC enters the cavity of pepsin, in which hydrophobic forces play a major role. The binding constant is 1.044 × 104 M−1 at 310 K. The upregulation or downregulation effect of THC on pepsin activity depends on its concentration. Molecular docking outcomes indicated that THC was encapsulated by various amino acids and established H-bonds with Tyr189 and Ser294, revealing that hydrogen bonds also contribute to maintaining the stability of THC-pepsin complex. In addition, the altered activity of pepsin may be related to the interaction between THC and the amino acids at the active site (Asp32) according to energy contribution results. 3D fluorescence spectroscopy, CD spectra and molecular dynamic simulations show that THC causes conformational changes in pepsin. The existence of THC makes pepsin structure to be less dense, leading to the decrease of energy traps. This suggests that pepsin becomes conformationally more suitable to bind to THC.

Daily rhythm in feeding behavior and digestive processes in totoaba (Totoaba macdonaldi) under commercial farming conditions

To identify daily changes in the digestive physiology of Totoaba macdonaldi, the feed intake, activity (pepsin, trypsin, chymotrypsin, lipase, amylase, and L-aminopeptidase), and gene expression (aminopeptidase and maltase-glucoamylase) of key digestive enzymes were measured in the intestine and the pyloric caeca. Fish were fed for three weeks every four hours during the light period to apparent satiation, and samples were taken every four hours throughout a 24-h cycle under a 12:12 L:D photoperiod. The feed consumption steadily increased until the third feeding (16:00 h, ZT-8) and decreased significantly towards the end of the day. The activity of pepsin and alkaline enzymes (trypsin, chymotrypsin, lipase, amylase, and L-aminopeptidase) exhibited a pattern dependent on the presence of feed, showing a significant reduction during the hours of darkness (ZT-12 to ZT-24). Expression of the intestinal brush border enzyme (L-aminopeptidase) increased during the darkness period in anticipation of the feed ingestion associated with the subsequent light period. The cosinor analysis used to estimate the feed rhythms for all tested enzymes showed that activity in the intestine and pyloric caeca exhibited significant rhythmicity (p < 0.05). However, no rhythmicity was observed in the intestinal expression of maltase-glucoamylase. Our results demonstrate that some of the behavioral and digestive physiology features of totoaba directly respond to rhythmicity in feeding, a finding that should be considered when establishing optimized feeding protocols.

Intragastric restructuring dictates the digestive kinetics of heat-set milk protein gels of contrasting textures

The gelation of milk proteins can be achieved by various means, enabling the development of diverse products. In this study, heat-set milk protein gels (15 % protein) of diverse textures were made by pH modulation and two gels were selected for dynamic in vitro gastric digestion: a spoonable soft gel (SG, pH 6.55′ Gʹ of ∼100 Pa) and a sliceable firm gel (FG, pH 5.65; Gʹ of ∼7000 Pa). The two gels displayed markedly different structural changes and digestion kinetics during gastric digestion. The SG underwent substantial structural compaction during the first 120 min of gastric digestion into a denser and firmer gastric chyme (26.3 % crude protein, G* of ∼8500 Pa) than the chyme of the FG (15.7 % crude protein, G* of ∼3000 Pa). These contrasting intragastric structural changes of the gels reversed their original textural differences, which led to slower digestion and gastric emptying of proteins from the SG compared with the FG. The different intragastric pH profiles during the digestion of the two gels likely played a key role by modulating the proteolytic activity and specificity (to κ-casein) of pepsin. Preferential early cleavage of κ-casein in SG stimulated coagulation and compaction of solid chyme, whereas rapid hydrolysis of αS- and β-caseins in the FG weakened coagulation. This study provided new insights into controlling the structural development of dairy-based foods during gastric digestion and modulating digestion kinetics.

Effects of Salinity on Growth, Digestive Enzyme Activity, and Antioxidant Capacity of Spotbanded Scat (Selenotoca multifasciata) Juveniles

As a euryhaline fish species that inhabits estuarine and coastal regions, the spotbanded scat (Selenotoca multifasciata) experiences growth influences during its larval stage due to variations in salinity. Here, we evaluated salinity required by early-stage spotbanded scat juveniles to achieve the highest growth performance, digestive enzyme activity, survival, and antioxidant capacity. We reared spotbanded scat juveniles (all 0.50 ± 0.05 g) in 0–35‰ salinity gradients for 50 days and recorded their survival rate every 10 days. After 50 experimental days, we measured morphological data, stomach and intestinal digestive enzyme activities, and liver antioxidant enzyme activities and malondialdehyde contents. In general, 5–15‰ salinity led to 100% survival. The 5‰ salinity group demonstrated the highest values for the following measures: final wet body weight; weight gain rate; specific growth rate; growth percentage; average daily gain; stomach amylase and lipase specific activities; and intestinal amylase, lipase, trypsin, and pepsin specific activities. However, stomach trypsin and pepsin activities did not demonstrate significant between-group differences (all p &gt; 0.05). The 25‰ salinity group demonstrated the highest liver superoxide dismutase and glutathione peroxidase activities and malondialdehyde content. Finally, the 0‰ salinity group demonstrated the highest liver catalase activity. Thus, spotbanded scat juveniles demonstrate the highest survival rates, growth performance, and digestive enzyme activity at 5‰ salinity and the strongest oxidative stress responses at 25‰ salinity.

Screening of Organic Acid Type and Dosage in Drinking Water for Young Rabbits.

Organic acids (OAs) are employed in animal feed to regulate gastrointestinal disorders and diarrhoea thanks to their ability to modulate the gastrointestinal environment and their antimicrobial capacity. However, there is not enough evidence regarding the most adequate OA and its effectiveness in rabbit farming. Therefore, the aim of this study was to screen and evaluate the response of young rabbits to six OAs, administered via drinking water, at three different concentrations (pH levels). Organic acids (acetic, ACET; formic, FOR; propionic, PROP; lactic, LAC; citric, CIT; and butyric, BUT) were tested at three concentrations (pH 3, 4, and 5). A negative control (CON; non-acidified water) was also included. We used 240 weaned rabbits (28 days old) divided into 2 batches. In each batch, animals were randomly allocated to 1 of the 19 experimental treatments and were housed in group cages of 6 animals per cage, treatment, and batch. Among the 240 rabbits, an additional cage with 6 animals was included to determine the initial physiological state of the animals. All animals were fed with commercial pelleted feed throughout the whole experiment. The duration of the study was 7 days, until 35 days of age. At 31 and 35 days of age, in each batch, three animals per day and treatment were slaughtered. The pH of the digestive contents in the fundus, antrum, duodenum, jejunum, ileum, and cecum, as well as the gastric pepsin enzyme activity, was measured. Water and feed consumption per cage and individual body weight (BW) were recorded daily. The type and dosage of OAs affected water intake. ACET 3, PROP 3, and BUT 3 reduced water intake compared to CON, negatively impacting feed intake and weight gain. FOR and CIT acids led to the highest BW and weight gain at 35 days, compared to PROP, LAC, and BUT (p < 0.05); showing ACET intermediate values. While OAs had limited effects on gastric and small intestine pH, acidified water at pH 4 and 5 lowered ileum and caecum pH (p < 0.05) compared to pH 3. Acidified water at pH 4 showed the highest (p < 0.05) pepsin activity compared to pH 3 and pH 5. Considering the limited sample size and short-term assessment period of our screening test, the OAs with the highest potential for use in post-weaning rabbits were FOR, ACET, and CIT at pH 4. The selected combinations did not exhibit any early adverse effects in young rabbits. These results should be further confirmed in a broader population of animals. It would also be advisable to extend the application of OAs over longer periods to evaluate their effects throughout the entire growing period of rabbits.

In vitro model to evaluate effect of acidic pepsin on vocal fold barrier function

The pathophysiology of laryngopharyngeal reflux (LPR) and its impact on the vocal fold is not well understood, but may involve acid damage to vocal fold barrier functions. Two different components encompass vocal fold barrier function: the mucus barrier and tight junctions. Mucus retained on epithelial microprojections protects the inside of the vocal fold by neutralizing acidic damage. Tight junctions control permeability between cells.Here we developed an in vitro experimental system to evaluate acidic injury and repair of vocal fold barrier functions. We first established an in vitro model of rat vocal fold epithelium that could survive at least one week after barrier function maturation. The model enabled repeated evaluation of the course of vocal fold repair processes. Then, an injury experiment was conducted in which vocal fold cells were exposed to a 5-min treatment with acidic pepsin that injured tight junctions and cell surface microprojections.Both of them healed within one day of injury. Comparing vocal fold cells treated with acid alone with cells treated with acidic pepsin showed that acidic pepsin had a stronger effect on intercellular permeability than acid alone, whereas pepsin had little effect on microprojections.This result suggests that the proteolytic action of pepsin has a larger effect on protein-based tight junctions than on phospholipids in microprojections. This experimental system could contribute to a better understanding of vocal fold repair processes after chemical or physical injuries, as well as voice problems due to LPR pathogenesis.

Network pharmacology combined with experimental validation reveals the mechanism of action of erpixing granules on functional dyspepsia

Ethnopharmacological relevanceFunctional dyspepsia (FD) is a prevalent gastrointestinal disorder characterised by high incidence and recurrence rates, posing significant health risks. Erpixing Granules (EPX), approved by the National Food and Drug Administration in 2002, are known for their spleen and stomach invigorating properties, effectively treating FD. However, its mechanism of action remains unclear. Aim of the studyThis study aims to elucidate EPX's mechanism of treating FD through network pharmacology, and experimental validation using FD animal models. MethodsIn this study, the chemical composition of EPX in positive and negative ion modes was analyzed by UHPLC-Q-TOF MS. The mass spectral data were processed and analyzed using MS-DIAL software to automatically match compound fragment information and identify the known components with the compound database to obtain the active components of EPX. SwissTargetPrediction was used to obtain EPX targets, while FD-related targets were sourced from GeneCards, OMIM and DisGeNET databases. A protein-protein interaction (PPI) network was constructed using the STRING platform, and potential signalling pathways of EPX were determined through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. Finally, an FD model was established in rates by administering a 0.1% iodoacetamide sucrose solution, followed by tail clamp stimulation to experimentally validate the network pharmacology findings. ResultsOur results revealed 139 effective ingredients in EPX, targeting 60 core FD-related genes. PPI network analysis identified EGFR, CTNNB1 and NFκB1 as core target genes. The KEGG pathway analysis indicated that EPX can modulate FD progression through the PI3K/AKT signalling pathway. Animal experiments demonstrated EPX's capacity to increase body mass, food intake and food utilisation efficiency in FD rats, alongside increased gastric juice secretion, pepsin activity, trypsin activity, cholesterol, bile acid and bilirubin activity. HE examination revealed that EPX improved the inflammatory infiltration of gastric mucosal cells in rats. Furthermore, EPX also promoted gastric emptying and intestinal propulsion in mice. These results suggest that EPX improves spleen and stomach function, enhances the protective effect on the spleen and stomach and promotes food digestion and absorption. Immunofluorescence studies revealed upregulated expression of PI3K, AKT and ANO1 proteins in gastric tissue following EPX administration, while Western blotting indicated increased expression of SCF and C-kit proteins. ConclusionSuggesting EPX's anti-FD effect may involve the regulation of the SCF/C-kit signalling pathway and activation of downstream PI3K/AKT signalling pathway, thereby promoting gastrointestinal motility and improving FD symptoms.

Long-Term Proton Pump Inhibitor-Acid Suppressive Treatment Can Cause Vitamin B12 Deficiency in Zollinger-Ellison Syndrome (ZES) Patients.

Whether the long-term treatment of patients with proton pump inhibitors (PPIs) with different diseases [GERD, Zollinger-Ellison syndrome (ZES), etc.] can result in vitamin B12 (VB12) deficiency is controversial. In this study, in 175 patients undergoing long-term ZES treatment with anti-acid therapies, drug-induced control acid secretory rates were correlated with the presence/absence of VB12 deficiency, determined by assessing serum VB12 levels, measurements of VB12 body stores (blood methylmalonic acid (MMA) and total homocysteine[tHYC]), and other features of ZES. After a mean of 10.2 yrs. of any acid treatment (5.6 yrs. with PPIs), 21% had VB12 deficiency with significantly lower serum and body VB12 levels (p < 0.0001). The presence of VB12 deficiency did not correlate with any feature of ZES but was associated with a 12-fold lower acid control rate, a 2-fold higher acid control pH (6.4 vs. 3.7), and acid control secretory rates below those required for the activation of pepsin (pH > 3.5). Over a 5-yr period, the patients with VB12 deficiency had a higher rate of achlorhydria (73% vs. 24%) and a lower rate of normal acid secretion (0% vs. 49%). In conclusion, in ZES patients, chronic long-term PPI treatment results in marked acid hyposecretion, resulting in decreased serum VB12 levels and decreased VB12-body stores, which can result in VB12 deficiency.

The effects of protein hydrolysates from rainbow trout by-products on growth, digestive and antioxidant enzymes, and liver lysozyme activity in sobaity (Sparidentex hasta) and Arabian yellowfin (Acanthopagrus arabicus) seabream juveniles

An eight-week feeding trial was conducted to elucidate the effects of rainbow trout by-products protein hydrolysates (FPH, 92 % peptides <1 kDa, ∼78.1 % crude protein, ∼2.7 % lipid) on growth and some selected physiological responses in sobaity (Sparidentex hasta) (1.7 ± 0.0 g) and Arabian yellowfin (Acanthopagrus arabicus) (0.9 ± 0.0 g) seabream. Fish meal in a basal diet (∼58 % crude protein, ∼12 % lipid) was replaced with FPH at four levels, including 0 (control), 1 % (FPH1), 2.5 % (FPH2.5), and 5 % (FPH5). Each experimental diet was offered thrice daily to fish in triplicate. At the end of the feeding trial, S. hasta fed with FPH diets showed positive linear and quadratic trends in growth performance, and those fed with FPH5 had the highest final weight (FBW; 9.8 ± 0.4) and better feed conversion ratio (FCR; 0.9 ± 0.0). Regarding A. arabicus it showed positive quadratic trend and fish fed FPH2.5 diet had the highest FBW (6.2 ± 0.3), and better FCR (1.2 ± 0.0) value. Trypsin and alkaline phosphatase (ALP) activities showed both positive linear and quadratic trends in S. hasta. In A. arabicus, both linear and quadratic trends were observed in the activities of trypsin and total proteases (P = 0.001), while positive quadratic trends in pepsin and ALP activities were noticed. Lysozyme activity significantly increased by supplementing the diet with FPH and showed both linear and quadratic trends in S. hasta, but it did not change in A. arabicus. Total antioxidant capacity (TAC) in A. arabicus increased and showed both linear and quadratic responses. The catalase and glutathione peroxidase activities in both species increased significantly in response to dietary FPH. This study showed that FPH effects in diet were species-specific and its supplementation levels at 2.5 and 5 % could be recommended in A. arabicus and S. hasta juveniles, respectively.

Pickering emulsion co-delivery system: Stimuli-responsive biomineralized particles act as particulate emulsifiers and bioactive carriers

Pickering emulsions provide a promising platform for the efficient delivery of bioactive. However, co-delivery of fragile bioactives with different physicochemical properties for comprehensive effects still faces practical challenges due to the limited protection for bioactives and the lack of stimuli-responsive property for on-demand release. Herein, a stimuli-responsive co-delivery system is developed based on biomineralized particles stabilized Pickering emulsions. In this tailor co-delivery system, hydrophilic bioactive (pepsin) with the fragile structure is encapsulated and immobilized by biomineralization, the obtained biomineralized particles (PPS@CaCO3) are further utilized as emulsifiers to form O/W Pickering emulsions, in which the hydrophobic oxidizable bioactive (curcumin) is stably trapped into the dispersed phase. The results show that two bioactives are successfully co-encapsulated in Pickering emulsions, and benefiting from the protection capacities of biomineralization and Pickering emulsions, the activity of pepsin and curcumin shows a 7.33-fold and 144.83-fold enhancement compared to the free state, respectively. Moreover, In vitro study demonstrates that Pickering emulsions enable to co-release of two bioactives with high activity retention by the acid-induced hydrolyzation of biomineralized particles. This work provides a powerful stimuli-responsive platform for the co-delivery of multiple bioactive compounds, enabling high activity of bioactives for the comprehensive health effects.

Investigating the impact of common migration substances found in milk packaging on proteases: A multispectral and molecular docking approach

The effects of common migration substances in milk packaging on digestive protease were studied. We choose the common migrants found in eight types of multi-layer composite milk packaging. Enzyme activity experiments revealed that pepsin activity decreased by approximately 18 % at 500 μg/mL of stearic acid and stearamide treatment, while trypsin activity decreased by approximately 18 % only by stearic acid treatment (500 μg/mL). Subsequently, fluorescence spectroscopy, circular dichroism spectroscopy, and molecular docking technology were employed to investigate the inhibition mechanism of protease activity by migrating substances in three systems: stearic acid-trypsin, stearic acid-pepsin, and stearamide-pepsin. Results showed that the inhibitory effect of stearic acid on trypsin is a reversible mixed inhibition, whereas the inhibitory effects of stearic acid and stearamide on pepsin are non-competitive. In all three systems, ΔH < 0, ΔS < 0, and ΔG < 0, indicating the binding process between the migrant and the protease is a spontaneous exothermic process primarily driven by hydrogen bonding and van der Waals forces. In addition, their binding constants are all around 104 L/moL, indicating that there are moderate binding affinities exist between migrants and proteases. The binding process results in the quenching of the protease's endogenous fluorescence and induces alterations in the enzyme's secondary structure. Synchronized fluorescence spectroscopy showed that stearic acid enhanced the hydrophobicity near the Tyr residue of trypsin. The molecular docking results indicated that the binding affinity of stearic acid-trypsin, stearic acid-pepsin, and stearamide-pepsin was –22.51 kJ/mol, −12.35 kJ/mol, −19.28 kJ/mol respectively, which consistent with the trend in the enzyme activity results. This study can provide references for the selection of milk packaging materials and the use of processing additives, ensuring food health and safety.

Effects of swimming activity and feed restriction on antioxidant and digestive enzymes in juvenile rainbow trout: Implications for nutritional and exercise strategies in aquaculture.

In this study, we investigated the effects of swimming activity and feed restriction on digestion and antioxidant enzyme activities in juvenile rainbow trout (average body weight of 26.54±0.36g). The stomach, liver and kidney tissues were obtained from four distinct groups: the static water group (fish were kept in static water and fed to satiation), the feeding restricted group (fish were kept in static water with a 25% feed restriction), the swimming exercised group (fish were forced to swimming at a flow rate of 1 Body Length per second (BL/s)) and the swimming exercised-feed restricted group (subjected to swimming exercise at a 1BL/s flow rate along with a 25% feed restriction). We determined the levels of glutathione, lipid peroxidation and the activities of catalase, superoxide dismutase, glutathione peroxidase, glutathione reductase, glucose-6-phosphate dehydrogenase and lactate dehydrogenase, as well as the presence of reactive oxygen species in the tissues obtained from the fish. Additionally, the activities of pepsin, protease, lipase and arginase in these tissues were measured. Swimming activity and feed restriction showed different effects on the enzyme activities of the fish in the experimental groups. It can be concluded that proper nutrition and exercise positively influence the antioxidant system and enzyme activities in fish, reducing the formation of free radicals. This situation is likely to contribute to the fish's development.