Effect of fat concentration on protein digestibility of Chinese sausage.

Chinese sausage is a popular traditional Chinese meat product, but its high-fat content makes consumers hesitant. The purpose of this study is to compare the nutritional differences of Chinese sausages with different fermentation times (0, 10, 20, 30 d) and fat content (the initial content was 11.59% and 20.14%) during digestion. The comparison of digestion degree, protein structure, and peptide composition between different sausages were studied through in vitro simulated digestion. Chinese sausages with high-fat content had higher α-helix, ß-turn, and random coil, making them easier to digest. The fermentation process made this phenomenon more pronounced. The high-fat sausage fermented for 10 d showed the highest release of primary amino acids (about 9.5%), which was about 3.5% higher than the low-fat sausage under the same conditions. The results of peptidomics confirmed the relevant conclusions. After gastric digestion, the types of peptides in the digestive fluid of high-fat sausages were generally more than those in low-fat sausages, while after intestinal digestion, the opposite results were observed. The type of peptide reached its peak after fermentation for 20 d. These findings are of obvious significance for selecting the appropriate fermentation time and fat content of Chinese sausages.

Effects of genetic variants and sialylation on in vitro digestibility of purified κ-casein.

Milk with different κ-casein (CN) phenotypes has previously been found to influence its gastric digestion rate. Therefore, the aim of the present study is to disentangle contributions of genetic variation and its related sialylation on the in vitro digestion process of κ-CN. Accordingly, κ-CN was purified from milk representing homozygous cows with κ-CN phenotypes AA, BB, or EE and used as substrate molecules in model studies using the INFOGEST 2.0 in vitro static digestion model. Furthermore, the effect of removal of the terminal sialic acids present on the O-linked oligosaccharides of the purified κ-CN A, B, and E protein variants were studied by desialylation enzymatic assays. The κ-CN proteins were purified by reducing anion exchange chromatography with purities of variants A, B, and E of 93.0, 97.1, and 90.0%, respectively. Protein degradations of native and desialylated κ-CN isolates in gastric and intestinal phases were investigated by sodium dodecyl sulfate-PAGE, degree of hydrolysis (DH), and liquid chromatography electrospray ionization mass spectrometry. It was shown that after purification, the κ-CN molecules reassembled into multimer states, which then constituted the basis for the digestion studies. As assessed by DH, purified variants A and E were found to exhibit faster in vitro digestion rates in both gastric and intestinal phases compared with variant B. Desialylation increased both gastric and intestinal digestion rates for all variants, as measured by DH. In the gastric phase, desialylation promoted digestion of variant B at a rate comparable with native variants A and E, whereas in the intestinal phase, desialylation of variant B promoted better digestion than native A or E. Taken together, the results confirm that low glycosylation degree of purified κ-CN promotes faster in vitro digestion rates, and that desialylation of the O-linked oligosaccharides further promotes digestion. This finding could be applied to produce dairy products with enhanced digestibility.

Phosphorylation and glycosylation isoforms of bovine κ-casein variant E in homozygous Swedish Red cow milk detected by liquid chromatography-electrospray ionization mass spectrometry.

Variations in the phosphorylation and glycosylation patterns of the common κ-casein (CN) variants A and B have been explored, whereas studies on variant E heterogeneity are scarce. This study reports for the first time the detailed phosphorylation and glycosylation pattern of the κ-CN variant E in comparison with variants A and B. Individual cow milk samples representing κ-CN genotype EE (n = 12) were obtained from Swedish Red cows, and the natural posttranslational modifications of its κ-CN were identified and quantified by liquid chromatography-electrospray mass spectrometry. In total, 12 unique isoform masses of κ-CN variant E were identified. In comparison, AA and BB milk consisted of 14 and 17 unique isoform masses, respectively. The most abundant κ-CN E isoform detected in the EE milk was the monophosphorylated, unglycosylated [1P 0G, ∼70%; where P indicates phosphorylation from single to triple phosphorylation (1-3P), and G indicates glycosylation from single to triple glycosylation (1-3G)] form, followed by diphosphorylated, unglycosylated (2P 0G, ∼12%) form, resembling known patterns from variants A and B. However, a clear distinction was the presence of the rare triphosphorylated, nonglycosylated (3P 0G, ∼0.05%) κ-CN isoform in the EE milk. All isoforms detected in variant E were phosphorylated, giving a phosphorylation degree of 100%. This is comparable with the phosphorylation degree of variants A and B, being also almost 100%, though with very small amounts of nonphosphorylated, glycosylated isoforms detected. The glycosylation degree of variant E was found to be around 17%, a bit higher than observed for variant B (around 14%), and higher than variant A (around 7%). Among glycosylation, the glycan e was the most common type identified for all 3 variants, followed by c/d (straight and branched chain trisaccharides, respectively), and b. In contrast to κ-CN variants A and B, no glycan of type a was found in variant E. Taken together, this study shows that the posttranslational modification pattern of variant E resembles that of known variants to a large extent, but with subtle differences.

Digestion Profiles of Protein in Edible Pork By-Products.

Edible pork by-products are widely consumed in many areas, whereas their digestion characteristics have rarely been evaluated. This work compared the digestibility of protein in boiled pork liver, heart, tripe and skin with tenderloin as a control. Cooked skin showed the highest digestibility in the simulated gastric digestion, whereas its gastric digests were less digested in the simulated intestinal stage. In contrast, cooked tripe showed the lowest gastric digestibility but relatively higher intestinal digestibility. All the edible by-products showed lower digestibility than tenderloin, especially for pork liver, in which large undigested fractions (>300 µm) could be observed. Corresponding to these results, larger amount of bigger peptides was found in the digests of pork liver and skin. In addition, peptides in tripe (average bioactive probability = 0.385) and liver digests (average bioactive probability = 0.386) showed higher average bioactive probability than other samples. Tripe digests contained the highest level of free Asp, Gln, Cys, Val, Phe, Pro, Ser, Thr, Ile and Asn, whereas heart digests contained the highest level of free Leu, Met and Arg. These results could help to reveal the nutrition value of pork by-products.

Differential in vitro digestion rates in gastric phase of bovine milk with different κ-casein phenotypes.

Casein (CN) micelles will coagulate in the stomach after ingestion, which is similar to the cheesemaking process. Although genetic variants of bovine proteins, especially κ-CN, have been confirmed to influence the coagulation properties of the CN micelle, its influence on milk digestibility has not been revealed yet. This study aimed to investigate how genetic variants, glycosylation degree of κ-CN, and CN micelle size influence digestion rates during in vitro gastrointestinal digestion. Three milk pools, representing κ-CN phenotypes of either AA, BB, or AB composition were prepared from milk of individual Danish Holstein cows representing these different genotypes. In vitro digestion of the 3 milk pools, AA, BB, or AB, was investigated by sodium dodecyl sulfate-PAGE, liquid chromatography-mass spectrometry, and degree of hydrolysis. The results showed that κ-CN AA milk had faster digestion rate in the gastric phase compared with BB and AB milks, whereas only small differences were apparent in the intestinal digestion phase. The results further documented that the milk pools representing κ-CN phenotypes BB and AB had comparable overall glycosylation degrees (50.9% and 50.0%, respectively) and higher than that of the AA milk pool (46.9%). Further, the AA milk pool was associated with larger CN micelles. These differences in CN micelle sizes and glycosylation degrees can be part of underlying explanations for the differential in vitro digestion rates observed between the AA, BB, and AB κ-CN milk pools.

Glycation from α-dicarbonyl compounds has different effects on the heat-induced aggregation of bovine serum albumin and ß-casein.

α-Dicarbonyl compounds are generated in large amounts during heat treatment in food production. This work compared the influence of glycation by α-dicarbonyl on the hydrothermal aggregation of bovine serum albumin (BSA) and of ß-casein (ß-CN). Glycation by α-dicarbonyl compounds was found to be more efficient than glycation by glucose in reducing the free amino groups, surface hydrophobicity and isoelectric point of BSA, thus greatly inhibited the hydrothermal aggregation of BSA. In addition, glycation by α-dicarbonyl greatly transformed the rigid BSA aggregates into flexible structures, based on analysis by fluorescence spectrum, transmission electron microscope and small-angle X-ray scattering. In contrast, both the aggregation process and aggregates conformation of ß-CN were found to be minimally affected by glycation, possibly due to the intrinsic disorder of ß-CN. This work highlights the substantial influences of α-dicarbonyl on dietary proteins during heat treatment depending on the protein structural characteristics.

Characterization and immunological activity of polysaccharides from Potentilla chinensis.

Three novel polysaccharides (PCPW, PCPS1 and PCPS2) were isolated from Potentilla chinensis and subjected to structural analysis by using spectral and physicochemical methods. The molecular weights of PCPW, PCPS1 and PCPS2 were calculated to be 4.45 × 103 Da, 1.18 × 104 Da and 4.23 × 104 Da, respectively. Analysis of monosaccharides composition confirmed that PCPW was composed of mannose, rhamnose, glucose, galactose and arabinose, while the two acidic polysaccharides PCPS1 and PCPS2 were consisted of six monosaccharides, including mannose, rhamnose, galacturonic acid, glucose, galactose and arabinose respectively. In addition, the main linkages of glycosidic bonds of PCPS2 were 1, 4-linked-rha, 1, 4-linked-man,1, 4-linked-galA and 1, 6-linked-man. Immunological tests indicated that both PCPW and PCPS2 could increase NO production of RAW264.7 cells, and promote splenocyte proliferation. All three polysaccharides proved to be activators of NF-κB. Overall, three polysaccharides showed a good immunological activity and pose great potential as a novel food or drug additive.

Peptidomic Investigation of the Interplay between Enzymatic Tenderization and the Digestibility of Beef Semimembranosus Proteins.

This work investigated the influence of enzymatic tenderization on digestibility changes of beef semimembranosus proteins using peptidomics methods. Hydrolysis by proteinase K and bromelain elevated the average bitterness index of identified peptides by generating high-Q values peptides (1714-1790 Cal/mol), including KDLFDPIIQ, LIDDHFLFDKPVSPL, and QLIDDHFLFDKPVSPLLL. Proteolysis during enzymatic tenderization acted as a "pre-digestion" step and significantly elevated the degree of hydrolysis of beef protein (by 4.5-17.3%) in subsequent simulated gastrointestinal digestion. Peptidomics analysis of digests revealed large variations in the peptide composition, which was positively correlated with the degree of proteolysis during enzymatic tenderization. Enzymatic tenderization with proteinase K- (for 0.5 h) or bromelain-treated samples largely increased the survival rate (by 65.5 or 82.8%) of peptides during simulated digestion, possibly because of the "secondary enzyme-substrate interaction" effect. This work could provide a new sight into the possible influence of enzymatic tenderization on meat nutrition.

Comparison of Free and Bound Advanced Glycation End Products in Food: A Review on the Possible Influence on Human Health.

Debate on the hazards of advanced glycation end products (AGEs) in food has continued for many years as a result of their uncertain bioavailability and ability to bind to their receptors (RAGEs) in vivo. There are increasing evidence that free and bound AGEs have many differences in gastrointestinal digestion, intestinal absorption, binding with RAGEs, in vivo circulation, and renal clearance. Therefore, this paper compares these aspects between free and bound AGEs by summarizing the available knowledge. On the basis of the current knowledge, we conclude that it is time to differentiate free AGEs from bound AGEs in food in future studies, because they vary in many aspects that are closely related to their influence on human health. Several perspectives were proposed at the end of this review for further exploring the difference between free and bound AGEs in food.

In Vitro Gastrointestinal Digestibility of Crystalline Oil-in-Water Emulsions: Influence of Fat Crystal Structure.

To investigate how the fat crystal structure affects lipid in vitro digestibility, 30% palm stearin-in-water emulsions were prepared after storage at different temperatures (4, 25, and 37 °C) for 1 h, which consisted of different polymorphic forms, sizes, and quantities of fat crystals. The variation of particle size ( d4,3), zeta potential, and microstructure during the gastrointestinal digestion and the free fatty acid (FFA) released in small intestine phase were investigated. After oral and gastric digestion, all of the emulsions underwent partial or complete coalescence and flocculation. During intestinal digestion, the d4,3 and zeta potentials did not notably affect lipid digestion. The FFA-released assay results indicated that the lipid digestion extent decreased as the fat crystal size and content of the ß polymorph increased, and there was no obvious relationship between FFA release and fat crystal quantity or solid fat content (SFC). This study highlighted the crucial roles of fat crystal size and polymorphic form in regulating the digestion behavior of lipid-based O/W emulsions.

Digestibility of glycated milk proteins and the peptidomics of their in vitro digests.

BACKGROUND: Milk proteins are widely used in food production and are often glycated by reducing sugar. Although many studies have reported the digestibility of glycated milk protein, most have focused on measuring degree of hydrolysis (DH), showing sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) image of digests. Detailed information on the changes in peptide composition of digests has seldom been revealed. Therefore, in addition to measuring the DH and showing the SGS-PAGE images of digests, we also analyzed the peptidomics in digests using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) and Mascot database in this work to further reveal the influence of glycation on protein nutrition. RESULTS: Compared with ß-lactoglobulin and bovine serum albumin (BSA), DH of ß-casein was suppressed to a lesser extent by glycation in both gastric and intestinal stages. Aggregates of glycated BSA were less sensitive to the action of digestive enzymes throughout gastrointestinal digestion according to SDS-PAGE images. Changes in the peptide composition of digests induced by glycation were distinctly displayed, showing both absence of peptides and occurrence of new peptides, based on the results obtained from LC-ESI-MS/MS. CONCLUSIONS: Glycation can greatly change the peptide composition in digests of milk protein. The nutritional impact of the change in the peptide composition requires further investigation, and the impact of MRPs in unabsorbed digests on the gut flora should be an interesting field for further studies. © 2018 Society of Chemical Industry.

Heat-induced amyloid-like aggregation of ß-lactoglobulin regulated by glycation: A comparison of five kinds of reducing saccharides.

The Maillard reaction occurs extensively in food industry and can change the aggregation behavior of dietary proteins. In this work, we reported the influence of glycation on the heat-induced amyloid-like aggregation behavior of ß-lactoglobulin (ß-Lg). Glycation was shown to accelerate the unfolding behavior of ß-Lg and thereby increased fibrillation rate during initial heating according to 1-anilinonaphthalene-8-sulfonate and thioflavin T assays. Prolongation of fibrils was found to be limited by glycation derived from glucose, fructose and lactose, possibly due to the decrease in ß-sheet structure of ß-Lg based on circular dichroism spectral results and transmission electron microscopy images. ß-Lg incubated with maltodextrin appeared to associate into globular micelles. Both covalent (disulfide bond and crosslinking structure) and non-covalent changes (steric hindrance and hydrogen bonds) induced by glycation are proposed to account for these fibrillation kinetic and conformational changes of ß-Lg aggregates. According to these findings, glycation are promising methods to regulate both fibrillation kinetics and fibril conformation in the food industry.

Digestibility of Bovine Serum Albumin and Peptidomics of the Digests: Effect of Glycation Derived from α-Dicarbonyl Compounds.

α-Dicarbonyl compounds, which are widely generated during sugar fragmentation and oil oxidation, are important precursors of advanced glycation end products (AGEs). In this study, the effect of glycation derived from glyoxal (GO), methylglyoxal (MGO) and diacetyl (DA) on the in vitro digestibility of bovine serum albumin (BSA) was investigated. Glycation from α-dicarbonyl compounds reduced digestibility of BSA in both gastric and intestinal stage of digestion according to measurement of degree of hydrolysis. Changes in peptide composition of digests induced by glycation were displayed, showing absence of peptides, occurrence of new peptides and formation of peptide-AGEs, based on the results obtained using liquid chromatography electron-spray-ionization tandem mass spectrometry (LC-ESI-MS/MS). Crosslinked glycation structures derived from DA largely reduced the sensitivity of glycated BSA towards digestive proteases based on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) results. Network structures were found to remain in the digests of glycated samples by transmission electron microscope (TEM), thus the impact of AGEs in unabsorbed digests on the gut flora should be an interest for further studies.

Physicochemical Properties and Chemical Stability of ß-Carotene Bilayer Emulsion Coated with Bovine Serum Albumin and Arabic Gum Compared to Monolayer Emulsions.

ß-carotene is a lipophilic micronutrient that is considered beneficial to human health. However, there are some limitations in utilizing ß-carotene in functional foods or dietary supplements currently because of its poor water dispersibility and chemical stability. A new type of ß-carotene bilayer emulsion delivery system was prepared by a layer-by-layer electrostatic deposition technique, for which were chosen bovine serum albumin (BSA) as the inner emulsifier and Arabic gum (GA) as the outer emulsifier. The physicochemical properties of bilayer emulsions were mainly characterized by droplet size distribution, zeta potential, rheological behavior, Creaming Index (CI), and encapsulation ratio of ß-carotene. Besides this, the effects of processing conditions (pH, thermal treatment, UV radiation, strong oxidant) and storage time on the chemical stability of bilayer emulsions were also evaluated. The bilayer emulsion had a small droplet size (221.27 ± 5.17 nm) and distribution (PDI = 0.23 ± 0.02), strong zeta potential (-30.37 ± 0.71 mV), good rheological behavior (with the highest viscosity that could reduce the possibility of flocculation) and physical stability (CI = 0), high ß-carotene encapsulation ratio (94.35 ± 0.71%), and low interfacial tension (40.81 ± 0.86 mN/m). It also obtained better chemical stability under different environmental stresses when compared with monolayer emulsions studied, because it had a dense and thick bilayer structure.