Enzymatic Cross-linking Of Bovine Gelatin As Well As Its Properties And Application

Gelatin is the denatured product of collagen, and consists of a heterogeneous mixture of water-soluble proteins of varying molecular weights. As one of the most important food proteins, gelatin is usually used in food processing as ingredient or additive. However, excessive hydrolysis of collagen is considered to limit the applications of gelatin. In order to further improve functional properties and expand the application field of gelatin, this study aimed to modify bovine gelatin via enzymatic cross-linking, and to obtain a cross-lined gelatin product with better properties.A ternary system comprising horseradish peroxidase(HRP), glucose oxidase and glucose was applied to cross-link bovine gelatin. Relative dityrosine content of the product was used as an index, to select suitable cross-linking conditions for gelatin via single factor trials. Electrophoretic analysis was used to confirm the formation of these macro-molecule polymers in the cross-linked gelatin. Physicochemical and functional properties of the cross-linked gelatin were assessed. After then, the cross-linked gelatin was applied to prepared edible film and set-style skimmed yoghurt, which aimed to to verify if the cross-linking was helpful to the properties of the film as well as quality of the yoghurt. The main results of this study were as follows.(1) Suitable conditions for gelatin cross-linkingIf protein concentration, p H and temperature of the reaction system were fixed at 50 g/kg, 7.0 and 37 °C, respectively, it was found that suitable HRP, glucose oxidase, glucose addition levels and reaction time were 100 U, 1 U, 0.025 mmol/g protein, and 3 h, respectively.(2) Physicochemical and functional properties of the cross-linked gelatinElectrophoretic analysis results demonstrated that the cross-linked gelatin contained much polymerized proteins. Comparing with the original gelatin, the cross-linked gelatin had the same amino acid compositions but more ordered secondary structure. The cross-linked gelatin showed enhanced thermal stability with higher decomposition temperature(323.7 °C versus 320.8 °C) but less mass loss(50.10% versus 50.90%). The cross-linked gelatin also had better colloidal stabilization in dispersion by forming aggregates with smaller hydrodynamic radius(73.3 versus 97.2 nm) and much negative zeta-potential(–8.15 versus –5.88 m V). Moreover, the cross-linked gelatin had increased surface hydrophobicity and emulsion stability by 39.2% and 54.7%, respectively, as well as greater emulsion stability during longer-storage period. However, the cross-linked gelatin had decreased emulsifying activity by 28.2%, and also showed decreased in vitro digestibility by 39.0% and 15.5% in pepsin hydrolysis and in pepsin-trypsin hydrolysis, respectively. The cross-linked gelatin exihibited increased gel strength by 12–14%, shortened gelation time by 20 min, increased gelling and melting temperature by 2 oC, respectively. The gels thus formed from the cross-linked glatin had higher elasticity modulus, which indicated that the cross-linked gelatin obtained better gelling properties.(3) Properties of the cross-linked gelatin filmComparing with the bovine gelatin film, the cross-linked gelatin film exhibited similar film thickness and moisture content, but lower water solubility(37.8% versus 49.9%) and higher opacity(0.71 versus 0.58. These results indicated that the cross-linked gelatin film had stronger water-resistence ability, and could prevent the illumination better. The cross-linked gelatin film also had enhanced tensile strength(32.0 versus 22.5 MPa), elongation at break(94.4% versus 81.1%), as well as higher melting point(86.9 °C versus. 80.8 °C), glass transition temperature(74.5 °C versus 72.3 °C) and storage modulus. These results indicated that the cross-linked gelatin film had improved mechanical properties and thermal stability. It was also evidenced that gelatin cross-linking resulted in 30.2% and 68.6% reductions in water vapor and CO2 permeability of the cross-linked gelatin film, respectively, but didn’t affect oil permeability. These results demonstrated that the cross-linked gelatin film had enhanced barrier properties. Furthermore, the cross-linked gelatin film possessed smaller cross-section voids(diameter 100?360 versus 200?595 nm). This fact supported that enzymatic cross-linking of gelatin could efficiently improve microstructure and properties of the gelatin film.(4) Quality attributes of the set-style skimmed yoghurtProtein, total sugar and total solids contents of the set-style skimmed yoghurts fortified with the bovine and cross-linked gelatin were about 32, 88 and 133 g/kg, respectively. When the storage time was prolonged, the two yoghurts had increased titration acidity and decreased p H values, but were detected to have similar textural indices. Comparing with the yoghurt added with gelatin, the yoghurt added with the cross-linked gelatin showed much decreased synereses(28.0% versus 31.5% on the first day; 7.0% versus 10.7% on the seventh day), higher viscosity, viscoelastic modulus and larger thixotropic loops area, together with more compact network structure. All these results suggested that the cross-linked gelatin could be used to improve the quality of the set-style skimmed yoghurt.