Study On The Preparation, Structures And The Biological Activities Of Egg White Protein Hydrolysates

China has been the largest country of egg production in the world. At present, however, the level of egg processing in China is very low and the production technology is obsolescent. Because of some dietary habits and the special needs of some industries, large consumption of egg yolk has been on demand, while egg white has been regarded as the waste, which is not only a waste of egg white resources, but a source of environmental pollution.Emzymatic hydrolysis of chicken egg white to produce egg white protein hydrolysates (EWPH) are investigated in this thesis, the contents are as following:Microplate reader was used to establish an in vitro method for the determination of antithrombin activity. The new method has some advantages, like low cost, high sensitivity and good repeatability.The contribution of the heat treatment of egg white before enzymatic hydrolysis to degree of hydrolsis (DH) was studied. The results showed that the heat treatment of egg white under acidic conditions did not significantly reduce the inhibition on the proteases, while the heat treatment under alkaline conditions could significantly eliminate the inhibition of egg white on the proteases. Circular dichroism measurement found the formation of aβ-sheet and random coil-based secondary structure in egg white proteins after heat treatment. The structural changes caused by the heat treatment of egg white proteins contributed little to the DH growth. The DH growth in egg white hydrolysis was ascribed to the elimination of inhibition of egg white on the proteases.DA201-C macroporous absorption resin (MAR) was chosen as the best desalination resin through the static absorption and the desorption experiments. The static adsorption rule of EWPH on DA201-C MAR was found. The investigation of the isosteric adsorption enthalpy led to the founding that adsorption of EWPH on DA201-C MAR was a physical adsorption process.By comparing the hydrolysis abilities of proteases, Alcalase and Protease N were chosen as enzymes with high capabilities to hydrolyze egg white proteins. The amino acid compositions of the hydrolysate from Protease N were similar to those from Alcalase. The amino acid scores of the hydrolysates of Alcalase and Protease N were higher than that of native egg white. The hydrolysates of Protease N had higher antithrombin activity and antioxidant activity than those of Alcalase. Protease N was eventually identified as the enzyme for egg white protein hydrolysates production.It was found that there existed significant relevance between the hydrophobic property and the antioxidant activity of EWPH. The molecular weight distribution of the fraction with the highest antioxidant activity was in the range of 249 to 4100 Da. The antithrombin activity of EWPH was closely related to the polarity and the charge nature of the amino acids in EWPH. After the gastrointestinal tract digestive enzymes treatment, EWPH reserved the antioxidant and the antithrombin activities well.Artificial neural network (ANN) was used to control and simulate the hydrolysis process of EMBR. The BP network could be used as a tool for simulating hydrolysis process. The EMRB product with highest antithrombin activity had an amino acid score of 113.25, with the first limiting amino acid of threonine. The free amino acid content was 0.39% (w/w). Two to eight amino-acid based peptides accounted for the relative content of nearly 80%.Four fractions with antithrombin activity were obtained by macroporous resin chromatography, gel filtration chromatography, semi-preparative RP-HPLC, analytical RP-HPLC. The amino acid sequences of 17 peptides were gained using MS/MS analysis. It was found that the peptides were rich in hydrophobic amino acids and positive charged amino acids. The peptide with the sequence of Leu-Val-Phe-Lys was syntheticly produced and the antithrombin activity was verified. The inhibition type of the bioactive peptide on thrombin was reversible. The theoretical analysis using molecular docking showed that after the docking the ligands matched very well with a water drainage region in the cavity of active site of thrombin. The increased hydrophobic side chains on peptide chain could help the increase of the dissolution and hydrophobic interaction in the process of molecular docking. Binding energy calculation showed that the increase of positive charge in peptide chain contributed to the formation of electrostatic match with the active cavity of thrombin, thus enhancing the binding force between the both.