| As the development of free radical theory and reactive oxygen species advance, “Antioxidant and Anti-aging” has become the hotspot in the cosmetic industry. Antioxidative peptides have huge developing potential in the field of cosmetics, because they are safe and stable, easily soluble in water, easy to absorb, with low molecular weight, and also can scavenge free radicals and chelate metal ion. Sheep placenta contains rich bioactive components and is the important protein resources. Although sheep are abundant in China, the utilization rate of sheep placenta is less than 3%. Sheep placenta by-product is the residue of sheep placent with plentiful protein after extracting the water-soluble bioactive peptide. Therefore, it is a high-quality protein resource to prepare peptides. In this paper, sheep placenta by-product was used as raw material to prepare antioxidative peptides. The peptides were purified, and in vivo experiments were conducted to study its anti-aging efficacy. The main contents were as follows:(1) The sheep placenta by-product powder was collected through 60-mesh screen after grinding the lyophilized precipitate that homogenized and centrifuged from sheep placenta. Its protein content was 81.41%, the polysaccharide content was 16.84%. The IC50 value of the sheep placenta by-product powder on scavenging DPPH radical, superoxide anion radicals and hydroxyl radicals was 5.4 mg/mL, 2.88 mg/mL and 15.2 mg/mL, respectively.(2) The DPPH radical scavenging activity(DRSA) and degree of hydrolysis(DH) were used as indexes, the preparation of sheep placenta hydrolysates(SPHs) were studied. The results showed that the enzymatic hydrolysates of papain possess the strongest scavenging activity of DPPH radical, and the DH was moderate. Accordingly, papain was adopted for hydrolysis. By single factor tests and response surface regression analysis, the optimum hydrolysis conditions were achieved as concentration of substrate 33 mg/mL, p H 6.4, temperature 55 oC, enzyme dosage 4900 U/g, and time 120 min. The resulted degree of DH was 7.92 %. Furthermore, SPHs prepared on this condition had strong scavenging activities of DPPH?, O2-?and ?OH, scavenging rate on them were 92.15%, 93.45% and 48.88% respectively(10 mg/mL).(3) The macroporous adsorption resins DA201-C were chosen for removing polysaccharides and desalination, and the removal rate of polysaccharides was 82.85%. After isolation by ultrafiltration, Sephadex G-10 gel filtration chromatography and RP-HPLC, the fraction(S12-2) with the highest DPPH? scavenging activities was obtained with the DPPH? scavenging rate of 32.52% at the concentration of 50 ug/mL. Finally, the structure and peptide sequence of fraction S12-2 was identified by MALDI-TOF-MS. The result showed it was a novel peptide composited of 6 amino acid residues with the peptide sequence of Glu-Pro-Val-Ser-His-Phe, and the molecular weight was 678.6.(4) The application in anti-aging cosmetics of fraction S12 obtained through gel column chromatography was studied. The S12 were added into skin care products, and the formulas and process of emulsion and cream containing different S12 content were designed. In double-blind test lasting 60 days, all cream and emulsion were significantly effective in improve skin hydration, reduce transepidermal water loss(TEWL) and improving skin elasticity. On the other hand, the sensory test of eye cream also suggested that the eye cream contained S12 has better senses making skin smooth, moisture-conditioned, fresh and soft. The eye cream containing sheep placenta antioxidant peptide can significantly fade out fine lines around eyes after using 60 days. |
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