Wuyin Weng

Stability of major allergen tropomyosin and other food proteins of mud crab (Scylla serrata) by in vitro gastrointestinal digestion

Stability in simulated gastric fluid is regarded as an important parameter for the estimation of food allergenicity. In this study, the digestive stability of allergenic protein tropomyosin (TM) and other food proteins from mud crab in simulated gastric fluid (SGF) and simulated intestinal fluid (SIF) digestion assay system was investigated and compared by SDS-PAGE and Western blot. In SGF system, proteins such as actin and the original band of myosin heavy chain (MHC) were rapidly degraded within a short period of time, while TM was relatively resistant to pepsin digestion. In SIF system, MHC was easily decomposed, while TM and actin were similarly resistant to digestion. Further study by IgE-immunoblotting and inhibition ELISA using sera from crab-allergic patients indicated that allergenicity of TM was partially decreased, suggesting proteinase digestion is effective in reducing the allergenicity of crab TM.

Effects of different processing methods on digestibility of Scylla paramamosain allergen (tropomyosin).

Crustacean allergy is a significant health problem around the world, and tropomyosin (TM) represents the major allergen of crustaceans. The aim of this study was to evaluate three processing methods (boiling, CUB, HPS) and identify the one method that is most effective in the degradation of TM and reduction of its IgE-binding reactivity, and make it easier to be decomposed during gastrointestinal digestion. SDS-PAGE analysis indicated that boiling had little impact on the digestive stability of TM. In contrast, combined ultrasound and boiling (CUB), and high pressure steaming (HPS) both could accelerate the digestion of TM. Similarly, western blotting and inhibition ELISA also demonstrated that the reactivity of IgG/IgE-binding of TM that was extracted from processed crab was partially decreased after treating with CUB or HPS. Among the three processing methods, HPS was the most effective method to accelerate the digestion of TM in gastrointestinal digestion, and reduce the reactivity of IgG/IgE-binding of TM. These results suggest that proper processing of crab could promote the degradation of TM in simulated gastrointestinal digestion, reduce the reactivity of IgG/IgE-binding of TM, and decrease the incidence of crab hypersensitivity in humans.

Effect of transglutaminase on properties of tilapia scale gelatin films incorporated with soy protein isolate

The effect of transglutaminase (TGase) on the properties of tilapia scale gelatin films in the presence of soy protein isolate (SPI) was investigated. When 3% TGase was added into gelatin films, the total soluble matter and protein solubility of films were decreased from 89.36% and 92.78% to 35.83% and 40.05%, respectively, and the decline was promoted by adding 5% SPI. The strength of the films was increased by adding 1% TGase irrespective of SPI addition, but decreased when the TGase concentration was further raised. No obvious colour change was observed in the films with or without TGase and SPI. Based on the results of SDS-PAGE, DSC and SEM, it was revealed that the movement of low molecular weight hydrophilic protein was depressed by the cross-linking network structure induced by TGase and SPI during film drying, indicating that adding SPI is essential to improve the thermal stability and water resistance properties of TGase-induced gelatin films.

Comparative study of in vitro digestibility of major allergen, tropomyosin and other proteins between Grass prawn (Penaeus monodon) and Pacific white shrimp (Litopenaeus vannamei).

BACKGROUND Stability in simulated gastric fluid is supposed to be an important parameter for the estimation of food allergenicity. In the present study, the digestive stability of allergenic protein tropomyosin (TM) and other food proteins from Grass prawn and Pacific white shrimp in simulated gastric fluid (SGF) and simulated intestinal fluid (SIF) digestion assay system was investigated and comparatively studied by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), western blotting, and inhibition enzyme-linked immunosorbent assay (ELISA). RESULTS In the SGF system, proteins such as actin and myosin heavy chain (MHC) were rapidly degraded within a short period of time, while TM was relatively resistant to pepsin digestion. In the SIF system, MHC was also easily decomposed, while TM and actin were resistant to digestion. Western blotting using a specific polyclonal antibody against TM indicated that the degradation pattern of shrimp TM by SGF and SIF was almost unaffected by the presence of other myofibrillar proteins. Further study by IgE immunoblotting and inhibition ELISA using sera from crustacean-allergic patients indicated that IgE binding of TM was decreased. CONCLUSION Proteinase digestion is effective in reducing IgE binding of shrimp TM. It is also of interest to notice that Pacific white shrimp TM had higher digestion stability than Grass prawn TM. However, Pacific white shrimp TM revealed enhanced IgE binding over that of TM from Grass prawn and thus it is possibly more allergenic.

Study on pepsinogens and pepsins from snakehead (Channa argus).

Three pepsinogens (PG1, PG2, and PG3) were highly purified from the stomach of freshwater fish snakehead (Channa argus) by ammonium sulfate fractionation, anion exchange, and gel filtration. Two-dimensional gel electrophoresis and native-PAGE analysis revealed that their molecular masses were 37, 38, and 36 kDa and their isoelectric points 4.8, 4.4, 4.0, respectively. All of the pepsinogens converted into their active form pepsins under pH 2.0 by one-step pathway or stepwise pathway. The three pepsins showed maximal activity at pH 3.0, 3.5, and 3.0 with optimum temperature at 45, 40, and 40 degrees C, respectively, using hemoglobin as substrate. All of the pepsins were completely inhibited by pepstatin A, a typical aspartic proteinase inhibitor. The N-terminal amino acid sequences of the three pepsinogens were determined to the 34th, 25th, and 28th amino acid residues, respectively. Western blot analysis of the three PGs exhibited different immunological reactions.

Comparative study of in vitro digestibility of major allergen tropomyosin and other food proteins of Chinese mitten crab (Eriocheir sinensis).

BACKGROUND China is the largest producer and consumer of aquatic products in the world; however, many people in China suffer from allergies upon consuming crab. Stability in simulated gastric fluid is regarded as an important parameter for the estimation of food allergenicity. RESULTS The digestive stability of allergenic protein tropomyosin (TM) and other food proteins from Chinese mitten crab in simulated gastric fluid (SGF) and simulated intestinal fluid (SIF) digestion assay systems was investigated and compared by SDS-PAGE, western blot and inhibition ELISA. In the SGF system, proteins such as the original band of myosin heavy chain (MHC) and actin were rapidly degraded within a short period of time, while TM was relatively resistant to pepsin digestion. In the SIF system, MHC was easily decomposed, while TM and actin were similarly resistant to digestion. Further study by IgE immunoblotting and inhibition ELISA using sera from crab-allergic patients indicated that allergenicity of TM was partially decreased. CONCLUSION Chinese mitten crab major allergen TM was resistant to pepsin while relatively susceptible to trypsin and chymotrypsin digestion. Both SDS-PAGE using purified TM and western blot using myofibrillar proteins indicated that the degradation pattern of TM by SGF and SIF was not affected by the presence of other myofibrillar proteins. Inhibition ELISA results revealed that proteinase digestion is effective in reducing the allergenicity of crab TM.

Separation and characterization of alpha-chain subunits from tilapia (Tilapia zillii) skin gelatin using ultrafiltration

Alpha-chain subunits were separated from tilapia skin gelatin using ultrafiltration, and the physicochemical properties of obtained subunits were investigated. As a result, α1-subunit and α2-subunit could be successfully separated by 100 kDa MWCO regenerated cellulose membranes and 150 kDa MWCO polyethersulfone membranes, respectively. Glycine was the most dominant amino acid in both α1-subunit and α2-subunit. However, the tyrosine content was higher in α2-subunit than in α1-subunit, resulting in strong absorption near 280 nm observed in the UV absorption spectrum. Based on the DSC analysis, it was found that the glass transition temperatures of gelatin, α1-subunit and α2-subunit were 136.48 °C, 126.77 °C and 119.43 °C, respectively. Moreover, the reduced viscosity and denaturation temperature of α1-subunit were higher than those of α2-subunit, and the reduced viscosity reached the highest when α-subunits were mixed with α1/α2 ratio of approximately 2, suggesting that α1-subunit plays a more important role in the thermostability of gelatin than α2-subunit.

Effect of blend ratio and pH on the physical properties of edible composite films prepared from silver carp surimi and skin gelatin

The effect of blend ratio and pH on the physical properties of surimi-gelatin composite films was investigated. Tensile strength (TS), film water solubility and soluble proteins of composite films increased with the increasing proportion of gelatin, while elongation at break (EAB) decreased. The TS of neutral films with the same ratio of surimi and gelatin were lowest, while increased at acidic or alkaline conditions. Similar tendency was also found in protein solubility and surface hydrophobicity of the film-forming solutions. On the other hand, the film water solubility and soluble proteins of neutral composite films were higher than those of acidic and alkaline films. Furthermore, it was revealed that the dissolved surimi and gelatin proteins could form strong composite films, which were insoluble in water. These results suggested that dissolved proteins were mainly involved in the formation of surimi-gelatin composite films.

Effect of setting temperature on glucono-δ-lactone-induced gelation of silver carp surimi

BACKGROUND Setting temperature is important for heat-induced surimi gel formation. However, there is little information concerning setting temperature on the properties of glucono-δ-lactone (GDL)-induced surimi gel, which is considered a new preparation technique. RESULTS The pH of surimi gel induced by 2% GDL was about 4.6, while the breaking force of GDL-induced surimi gel preheated at a temperature range of 35-50 °C was higher than that of heat-induced surimi gel. The breaking force, deformation and whiteness of GDL-induced surimi gel were increased with increasing setting temperature from 30 to 45 °C, but water-holding capacity was decreased. When setting temperature was further increased to 50 °C, the textural properties were decreased, and myosin heavy chain (MHC) was degraded slightly. The data of protein subunits solubilized in various solvents revealed that MHC participated in the formation of GDL-induced surimi gel mainly through hydrophobic interactions. Furthermore, when GDL-induced surimi gel was preheated at 45 °C, a compact and fine fiber microstructure was observed by scanning electron microscopy. CONCLUSION Setting treatment at the appropriate temperature could promote the formation of a fine, compact GDL-induced surimi gel network, resulting in improved textural properties.

Preparation and bioavailability of calcium-chelating peptide complex from tilapia skin hydrolysates

BACKGROUND With the continuous improvement in material life, the generation of fish by-products and the market demand for calcium supplements have been increasing in China. Therefore a calcium-chelating peptide complex (CPC) from tilapia skins was prepared and its effect on calcium (Ca)-deficient mice was investigated. RESULTS The molecular weight distribution of CPC mainly ranged from 2000 to 180 Da, and its contents of complete amino acids and free amino acids were 85.30 and 8.67% (w/w) respectively. Scanning electron microscopy images and Fourier transform infrared data revealed that Ca crystals were bound with gelatin hydrolysates via interaction between Ca ions and NH/CN groups. When Ca-deficient mice were fed CPC and CaCO3 respectively for 4 weeks, no significant differences in serum biochemistry or bone mineral density were found. However, the length, weight, Ca content and hydroxyproline content of the femur, Ca absorption and body weight gain of mice fed CPC were significantly higher than those of mice fed CaCO3 . CONCLUSION It is concluded that the prepared CPC could promote bone formation via better bioavailability of Ca and an increase in bone collagen.