Preparation Of Chicken Feets Skin Collagen Polypeptide And Research On Its Inhibition Of Tyrosinase

Collagen Peptide has been a kind of extensively studied peptides Currently, using enzymaticproduction of bioactive collagen peptides, is not only conductive taken full advantage of the richresources of collagen, but also has a wide range of dietary and pharmaceutical prospects. Because ofits safety, functional diversity, Collagen peptides has been gradually recognized; There are manypolypeptide products with lower blood pressure, anti-oxidation, nutritional adjustment and otherfunctions have already appeared on the market. In this paper, chicken feet skin was used as rawmaterials, and using response surface mathematical analysis to optimize the extraction process ofchicken collagen peptides, then preliminary isolated and purified the collagen peptides,studied itstyrosinase inhibitiory activity function. Finally, using B16melanoma cells to verify the security ofchicken collagen peptides and the effect on the synthesis of melanin.Firstly, The preparation of chicken collagen peptides were optimized. Degree of hydrolysis(DH)as indexes, screened alkaline protease from seven commercial enzymes as an experimental enzyme.Then single factor experiments with temperature(T), pH, concentration of substrate(S), the amount ofenzyme (E), the hydrolysis time, five factors have done to optimize enzyme solution; Select the threefactors that affect digestion significantly, DH as the response value, using the response surfaceanalysis with three factor and three level. Research on effects of temperature (T), pH, substrateconcentration (S) on the digestion process. The optimum parameters of the producing chicken feetcollagen polypeptide were temperature61℃, pH8.10, substrate concentration5.10%, plus theamount of enzyme2500U/g, enzymolysis time of5h, the maximum degree of hydrolysis was17.04±0.26%.Using ultrafiltration to grade chicken feet collagen polypeptide which has been prepared byoptimal hydrolysis conditions. Taking tyrosinase inhibition rate as an index to screening molecularweight less than3kd polypeptide（components III）as experiment objective. Sample concentration onultrafiltration and ultrafiltration feed inlet pressure were optimized to get the best conditions for theUF: sample concentration of1.5%, ultrafiltration pressure of200Kpa. The components III weredesalted by macroporous resin adsorption. Peptide recovery rate as the index to optimize the sampleflow rate and ethanol concentration, ethanol volume fraction, the optimal results are obtained as follows: the sample concentration of35mg·mL-1, ethanol flow rate of90mL·h-1, ethanol volumefraction of70%, The peptide recovery was96.70±0.76%.The components III were freeze-drying after desalted, then performed the inhibition of tyrosinaseactivity, and its tyrosinase inhibition kinetics. Measured IC50values of chicken collagen peptidesinhibition on the tyrosine monophenolase’s enzyme and the tyrosinase diphenols’ enzyme, whichwere0.570mg mL-1and0.426mg·mL-1.Tyrosine diphenols are reversible inhibition ofnoncompetitive inhibition, Michaelis constant Km and inhibition constants were24.68μmol·mL-1and10.76mg·mL-1.With B16melanoma cells as a model, exploring the safety of chicken collagen peptides and theirimpact on melanin synthesis. The results showed that the chicken feet collagen polypeptideconcentration in the range of0to200μg/mL has no effect on the proliferation of cells, and caneffectively inhibit the formation of melanin,200μg·mL-1collagen peptide concentrations, melaninrelative content reduced to57.83%.