Abstract: In this study, the effect of dual-enzyme hydrolysis on the allergenicity of the major proteins in skimmed milk was evaluated based on the degree of hydrolysis, molecular mass distribution, immunoglobulin G (IgG) binding capacity and allergic patients’ serum IgE binding capacity of hydrolysates. The results showed that after one-step hydrolysis for 80 min at an enzyme/substrate ratio of 3 000 U/g and an Alcalase to Flavourzyme ratio of 3:1, the IgG binding capacity of casein, α-lactalbumin and β-lactoglobulinin in skimmed milk decreased significantly by 74.93%, 97.24% and 93.46%, respectively (P < 0.05).The advanced structure of proteins was damaged, leading to an increase in the α-helical and β-turn contents, and a decrease in the β-sheet content. In addition, the free sulfhydryl content increased significantly to 14.29 μmol/g (P < 0.05), the surface hydrophobicity also increased, and the structure became loose. The particles in skimmed milk were aggregated, causing the average particle size to increase significantly to 346.90 nm (P < 0.05) and reducing the stability of skimmed milk. The binding capacity to serum IgE from patients with milk allergy of the enzymatic hydrolysate decreased significantly by 23.53% (P < 0.05), and the overall allergenicity decreased significantly by 69.60% (P < 0.05). Therefore, dual-enzyme hydrolysis is an effective method to reduce the allergenicity of skimmed milk.

Key words: skimmed milk; protease; allergenicity; enzyme-linked immunosorbent assay

摘要： 以水解度、分子质量分布及酶解产物对免疫球蛋白G（immunoglobulin G，IgG）和患者血清IgE结合能力为 指标，采用双酶水解方法研究酶对脱脂牛乳体系主要蛋白致敏性的影响。结果表明：双酶一步水解E/S为3 000 U/g、 碱性蛋白酶与风味蛋白酶配比为3∶1、酶解80 min条件下脱脂牛乳中主要致敏乳蛋白酪蛋白、α-乳白蛋白、β-乳球蛋 白的IgG结合能力分别显著下降74.93%、97.24%、93.46%（P＜0.05）；酶解破坏脱脂牛乳蛋白高级结构，使α-螺 旋、β-转角结构增加，β-折叠结构减少，游离巯基含量显著增加至14.29 μmol/g（P＜0.05），表面疏水性增强，结 构趋于松散；脱脂牛乳体系中粒子发生聚集，平均粒径显著增加至346.90 nm（P＜0.05），稳定性降低；酶解产物 对牛乳蛋白过敏患者血清IgE结合能力显著降低23.53%（P＜0.05），综合致敏性显著下降69.60%（P＜0.05）。因 此双酶水解是降低脱脂牛乳致敏性的有效方法。

关键词: 脱脂牛乳；蛋白酶；致敏性；酶联免疫吸附测定