紫苏籽粕蛋白源抗氧化肽的纯化、结构鉴定及体外抗氧化活性

通过单因素和响应面实验对紫苏籽粕蛋白的闪提工艺进行了优化,将所获蛋白通过碱性蛋白酶酶解,酶解物依次通过超滤和柱层析进行分离纯化,并对抗氧化活性最高的多肽进行液质分析。结果表明,紫苏籽粕蛋白提取的最优工艺参数为：料液比1︰20(g/mL)、闪提pH 10、闪提时间30 s和闪提转速3000 rpm。蛋白酶解物经超滤分离后,其分子量小于3kDa的组分具有较高的抗氧化活性、该组分经DEAE-32离子交换色谱分离后, 得到D1、D2、D3共3个组分,其中D1的抗氧化活性最好,D1经Sephadex G-25凝胶层析分离纯化后,富集得到G1和G2两个组分,组分G1抗氧化活性最高。抗氧化肽G1对DPPH、ABTS、O_2^-、.OH 自由基清除率及还原力依次为98.97%、85.11%、91.83%、93.40%、0.477；通过LC-MS-MS鉴定,抗氧化活性肽G1组分主要由15个氨基酸组成,荷质比m/z为672.39,分子质量为1718.82 Da,其氨基酸序列为Glu－Met－Pro－Tur－lle－Ala－Ser－Met－Gly－lle－Tyr－Val－Val－Ser－Lys。

Purification and Structural Identification of Antioxidant Peptide from Perilla seed meal Protein and Antioxidant Activity in Vitro

The flash extraction process of perilla seed meal protein was optimized through single factor and response surface experiments. The protein was hydrolyzed by alkaline protease, and the hydrolysates were separated and purified by ultrafiltration and column chromatography. The peptides with the highest antioxidant activity were analyzed by liquid chromatography-mass spectrometry. The results showed that the optimum extraction parameters were as follows: material-liquid ratio 1:20 (g/mL), flash-extraction pH10, flash-extraction 30s and flash-extraction speed 3000 rpm. The proteolysis product was then separated by ultrafiltration, and the small-sized fraction (<3 kDa) possessed higher antioxidant activities. This fraction was further separated into three fractions (D1, D2, D3) by DEAE-32 ion exchange chromatography. It was found that fraction D1 possessed higher antioxidant activities. Finally, the fraction D1 was separated into two fractions (G1, G2) by Sephadex G-25 gel chromatography, the antioxidant activity of the fraction G1 was the highest. The scavenging rates of the antioxidant components G1 to DPPH, ABTS, O2_^- and OH were 98.97%, 85.11%, 91.83%, 93.40%, respectively, and the reducing power of G1 was 0.477. By LC-MS-MS identification,the charge-to-mass ratio m/z is 672.39, and the molecular mass is 1718.82 Da, and its amino acid sequence was Glu-Met-Pro-Tur-lle-Ala-Ser-Met-Gly-lle-Tyr-Val-Val-Ser-Lys.