AB-8大孔树脂-C18柱联用分离干腌牛肉抗氧化肽的研究

Isolation of antioxidant peptides from dry corned beef by AB-8 macroporous resin-C18 column

该文利用盐酸提取法提取了干腌牛肉多肽,借助AB-8型大孔树脂及C18球形硅胶柱的分离纯化出4种抗氧化肽,利用分子对接模拟了4种抗氧化肽与超氧化物歧化酶(superoxide dismutase,SOD)的结合位点。利用盐酸提取法提取了抗氧化肽并测试其在1、2、3、4、5 mg/mL的抗氧化能力,发现在5 mg/mL时,DPPH自由基、ABTS阳离子自由基和超氧阴离子自由基(·O_(2)^(-))清除率分别为86.24%、35.62%和54.34%。对比了AB-8、D101、S-8、X-S、HPD-500、NKA-9的吸附率与解吸率,AB-8的吸附率与解吸率最好,分别为92.55%和82.88%。研究优化了AB-8的上样流速、上样量、乙醇洗脱浓度和洗脱流速,在4 BV/h上样流速、39 mL上样量、75%(体积分数)乙醇溶液洗脱剂和1 mL/min洗脱流速时,可以分离出3个组分(A、B和C),测试3个组分的抗氧化能力,B组分的DPPH自由基、ABTS阳离子自由基和·O_(2)^(-)清除能力最高,分别为73.33%、21.01%和63.33%。对B组分进行C18柱分离,分离出4个抗氧化肽:FDGDF、TGPGPW、FLSDH和KPFDAK。与SOD(PDB ID;1E9O)进行分子对接,发现FDGDF、TGPGPW、FLSDH和KPFDAK均可以与SOD形成氢键,结合能分别为-6.7、-6.55、-6.6、-7.35 kcal/mol,1 mg/mL的FDGDF可以有效增加SOD活力11.64%,酶活力单位增加697 U/mL,研究结果为干腌牛肉制品的开发提供理论基础。

The dry corned beef polypeptides were extracted by hydrochloric acid extraction method.Four kinds of antioxidant peptides were isolated and purified by AB-8 macroporous resin and C18 spherical silica gel column.The binding sites of four kinds of antioxidant peptides and superoxide dismutase(SOD)were simulated by molecular docking.The antioxidant peptides were extracted by hydrochloric acid extraction method and their antioxidant capacity was tested at 1,2,3,4,and 5 mg/mL.The scavenging rates of DPPH free radicals,ABTS cationic radicals,and superoxide anion radicals were 86.24%,35.62%,and 54.34%,respectively,at 5 mg/mL.The adsorption rate and desorption rate of AB-8,D101,S-8,X-S,HPD-500,and NKA-9 were compared.The adsorption rate and desorption rate of AB-8 were the best,which were 92.55%and 82.88%,respectively.The loading velocity,loading volume,ethanol elution concentration,and elution velocity of AB-8 were optimized,which were 4 BV/h of loading velocity,39 mL of loading velocity,75%(volume fraction)of ethanol solution elution agent,and 1 mL/min of elution velocity,respectively.Three components(A,B,and C)could be separated to test the antioxidant capacity of the three components.The scavenging capacity of DPPH free radicals,ABTS cationic radicals,and super-oxide anion radicals of component B was the highest,which were 73.33%,21.01%,and 63.33%,respectively.Four antioxidant pep-tides were isolated by C18 column separation of component B,including FDGDF,TGPGPW,FLSDH,and KPFDAK.Through molecular docking with SOD(PDB ID;1E9O),it was found that FDGDF,TGPGPW,FLSDH,and KPFDAK could form hydrogen bonds with SOD,and the binding energies were-6.7,-6.55,-6.6,and-7.35 kcal/mol,respectively.1 mg/mL of FDGDF could effectively increase SOD activity by 11.64%,and enzyme activity unit by 697 U/mL.It provides the theoretical basis for the development of dry corned beef products.