Study On The Preparation, Purification And Identification Of Antioxidant Peptides From Se-enriched Tea Protein

Selenium is an important trace elements in the nutrition of plants, animals and microorganism, its biological function is mainly presentsd by selenoproteins and peptides containing selenium. Tea protein has a certain antioxidant activity. Compared with other biological molecules, the biological activity and diversity of peptides, whose molecular weight between protein and amino acid, is better. What’s more, the antioxidant activity and security of peptides is higher. It will have great application value to efficiently band the study on selenium and tea protein peptides together to explor the activity of se-contained tea protein peptides.In this paper, se-enriched alkali soluble tea protein, extracted from se-enriched tea, were hydrolyzed by different proteases to get peptides with higher antioxidant activity. And purification and identification of antioxidant peptides were studied.The main content of this paper were the following parts:(1) Compared with the general tea, polyphenols in se-enriched tea decreased and free amino acids increased. So selenium may improve the taste of tea through reducing the astringency and improving freshness. At the same time, the content of organic selenium in se-enriched tea were more than 80%, which conformed to the standards of the government.(2) The mass ratio of tea polyphenols and tea protein as index, the optimal process parameters were obtained through the typical analysis and ridge analysis as follows: PVPP of 4.37 mg/mL, NaOH of 0.14 M, temperature of 30 ℃.In this condition, the mass ratio of tea polyphenols and tea protein were 0.089.(3) The ability of scavenging free radicals as index, Papain was chosen as the best enzyme in preparing antioxidant peptides from the tested four proteases. The optimal process parameters were obtained through the typical analysis and ridge analysis as follows: pH of 7.45,temperature of 55 ℃,enzyme of 1900 U/g,time of 3.4 h. In this condition, the hydroxyl free radical inhibition rate of the antioxidant peptides was 66.9%.(4) Three fractions, TPepⅠ(<1 kDa), TPepⅡ(1~10 kDa) and TPepⅢ(10~100 kDa), were got from se-enrich alkali soluble tea protein peptide mixture through ultrafiltration, and the order of the ability of scavenging free radicals was that TPepⅠ> TPepⅢ> TPepⅡ, but the order of selenium content was that TPepⅢ>TPepⅡ>TPepⅠ. Fraction TPepⅠwas chosen to be purified by Sephadex G-25 further, two fractions, TPep Ⅰ-1 and TPep Ⅰ-2, were obtained, and their ability of scavenging free radicals were that TPepⅠ-1>TPepⅠ-2, as same as their selenium content. Chemical bonds of N-H, O-H, C-H, C=O and C-N were found in TPepⅠ-1 by IR, and two antioxidant peptides sequence, LPMFG(Leu-Pro-Met-Phe-Gly) and YPQSFIR(Tyr-Pro-Gln-Ser-Phe-Ile-Arg), were got from TPep Ⅰ-1 through automatic Amino Acid Analyzer and UPLC-Q-TOF-MS. The molecular weight of those peptides were 563.27 Da and 909.47 Da. The antioxidation mechanism may be that: 1) Hydrophobic amino acids could combine with oxygen or inhibit the release of hydrogen in the lipid to slow down the chain reaction of lipid peroxidation; 2) Pro and Phe could chelate with metal ions to terminate the redox reaction; 3) Leu and Tyr could provide protons, which can combine with free radicals; 4) The condensation products of Ser, Gly and salicylic aldehyde could chelate with Fe2+ to inhibit the generation of hydroxyl radicals.