Bioactive polypeptide LVYPFPG as well as preparation method and application thereof
A technology of biologically active peptides and derivatives, applied in the field of proteins, can solve problems such as human risks, and achieve the effects of reducing injuries, regulating the body's immune activity, and enhancing antioxidant activity
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[0033] Example 1 Artificial synthesis of active peptide LVYPFPG
[0034] 1. Synthesis of bioactive peptides
[0035] 1. Weigh 3g of RINK resin (degree of substitution 0.3mmol / g) into a 150ml reactor and soak it with 50ml of dichloromethane (DCM).
[0036] After 2.2 hours, the resin was washed with 3 times the resin volume of nitrogen-dimethylformamide (DMF), and then drained. This was repeated four times, and the resin was drained before use.
[0037] 3. Add a certain amount of 20% piperidine (piperidine / DMF=1:4, v:v) to the reactor, and shake it on a decolorizing shaker for 20 minutes to remove the Fmoc protecting group on the resin. After removing the protection, wash it four times with 3 times the resin volume of DMF, and then drain it.
[0038] 4. Take a small amount of resin and use the ninhydrin (Ninewell Ninhydrin) method to detect (two drops each for inspection A and inspection B, reaction at 100°C for 1 min), the resin is colored, indicating successful deprotection.
[0039] 5....
Example Embodiment
[0082] Example 2 Antioxidant activity experiment of biologically active peptide
[0083] The free radical scavenging method (DPPH·method) and the total antioxidant capacity method (Ferric Reducing AbilityPower FRAP method) were used to test the antioxidant activity of the biologically active polypeptide LVYPFPG obtained in Example 1.
[0084] 1. [DPPH·] method to determine the in vitro antioxidant activity of the biologically active peptide LVYPFPG
[0085] 1) Experimental reagents and instruments
[0086] Reagents: 1,1-Diphenyl-2-picrylhydrazyl (1,1-Diphenyl-2-picrylhydrazyl[DPPH·]), produced by Wako, Japan; methanol, provided by Shanghai Sinopharm; obtained in Example 1 Milk-derived biologically active peptide LVYPFPG.
[0087] Main instruments: Sunrise microplate reader, product of Tecan, Austria; 96-well cell culture plate, manufactured by Millipore, USA; analytical balance, product of Meitelei-tolido.
[0088] 2) Experimental method
[0089] (1) 1mmol / L[DPPH·] methanol solution
[00...
Example Embodiment
[0122] Example 3 Experiments on the activity of biologically active peptides to promote immunity
[0123] 1. The in vitro lymphocyte proliferation ability of the biologically active polypeptide LVYPFPG determined by MTT method
[0124] 1) Experimental materials and instruments:
[0125] Reagents and materials: experimental animal balb / c mice (male 6-8 weeks old, Animal Experiment Center, School of Agriculture and Biology, Shanghai Jiaotong University); milk-derived biologically active polypeptide LVYPFPG obtained in Example 1; mouse lymphocyte extract (Purchased from Soleibao); RPMI1640 medium (purchased from GIBCO); 3-(4,5-dimethylthiazole-2)-2,5-diphenyltetrazolium bromide (MTT, purchased From Amresco); Concanavalin (ConA, purchased from Sigma); Bovine Serum Albumin (BSA, purchased from Genebase); Pepsin (purchased from Sigma); Pancreatin (Corolase PP, purchased from AB) ).
[0126] Equipment: LRH-250F biochemical incubator, Shanghai Heng Technology Co., Ltd.; GL-22M high-speed re...
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