The Preparation Of Enzymatic Hydrolysis Egg-white Peptides And Its Application In Cell Culture

The enzymatic hydrolysis egg-white peptides have antioxidant,antibacterial,ant-fatigue and other effects.The products of egg-white peptides had been widely applied in the field of food health field.Bioactive peptides have the effect of promoting cell growth.However,the egg-white peptides application in cell culture is rarely reported.This research utilized high-pressure technique to enzymatic hydrolyzed egg-white protein.The egg white peptides were successfully prepared through optimizing process.The experimental study of the article includes basic physical and chemical properties and on the growth of adherent cells.The result provided the theoretical basis for the application of egg white peptides in cell culture.First,the preparation methods and research status of biologically active peptide,protein hydrolyzate and egg white hydrolyzate were introduced.Secondly,the process of egg-white protein proteolysis was optimized by enzyme screening(Alkaline protease,Papain,Pancreatin,Neutral protease,Pepsin,Trypsin),single factor experiment and orthogonal experiments with the degree of hydrolysis indicator.The results showed that the hydrolysis of trypsin was better than other five enzymes in egg-white protein proteolysis.The optimum hydrolysis was:120MPa,50?,pH7.0 and enzyme/substrate(E/S)?1:2.5(mass ratio).The degree of hydrolysis could reach to 45.13%.Then,the physical and chemical properties of egg-white protein proteolysis were studied.The results showed that the high pressure enzymolysis of egg-white peptides had high solubility,thermal stability and foaming property at pH 2-10,and the stability of foaming was low.Amino acid detection and molecular weight detection proved that it has a high degree of hydrolysis and contain a large number of amino acids which were beneficial to cell growth.Finally,the paper used BHK-21 and Marc-145 cells to detect the possibility of the high pressure enzymolysis of egg-white peptides application in cell culture,including sub culturing,growth curve test,serum reduction test,and separation and purification methods of cellulose media.The results showed that the maximum amount of proliferation of growth curve was 6.69 X 10~5 cells/mL and 4.75 ×10~5 cells/mL,which was 1.56 and 1.73 times of the control group.The cell doubling time was 21.25h and 27.94h,which reduced 6.97h and 4.24h compared with the control group.The two kinds of cells could be stable for more than 15 generations,and the growth morphology did not change significantly.The experiment also showed that the high pressure enzymolysis of egg-white peptides could reduce the serum content in cell culture,when it addition was 0.25%.Isolation and purification experiments showed that the self-made egg-white peptides might contain the substance that inhibits cell growth--EH4.If EH4 is removed from the high pressure enzymolysis of egg-white peptides,whether or not its cell growth is more visible and needs further study.In conclusion,the egg-white peptides,which utilized high-pressure technique to enzymatic hydrolyzed egg-white protein,had the activity of promoting cell growth.It has application prospect and development value in large-scale cell culture.