Abstract: To optimize the ultrasound⁃assisted enzymatic extraction process of selenium proteins from selenium⁃enriched Lentinula edodes and systematically evaluate their nutritional quality and antioxidant activity，single⁃factor experiments were carried out using protein extraction rate and total selenium extraction rate as response indicators to determine the parameter range. The Box⁃Behnken response surface methodology was then adopted to establish a regression model for extraction process optimization.Subsequently，the optimized process was applied to four groups of Lentinula edodes raw materials with different total selenium contents（S0：0.3±0. 0 mg/kg；S1：9.8±0.4 mg/kg；S2：20.0±0.9 mg/kg；S3：99. 6±1. 1 mg/kg）. The selenium speciation，amino acid compositions and in vitro antioxidant capacity of selenium protein extracts obtained from different raw materials were compared and analyzed.The results of process optimization showed that the optimal conditions were as follows：alkaline cellulase addition amount of 0.6%（mass ratio），solid⁃liquid ratio of 1∶25（g/mL），pH value of 9.0，enzymolysis temperature of 48 ℃，ultrasonic power of 300 W，and ultrasonic time of 54 min. Under these conditions，the protein extraction rate predicted by the model could reach 50. 02%. Selenium speciation analysis indicated that selenoamino acids accounted for up to 86.6% of the total selenium in the selenium protein extract of S3，with selenocysteine as the predominant existing form. Amino acid nutritional evaluation revealed that the S3 selenium protein extract exhibited superior values in the ratio of essential amino acids to total amino acids（40.32%），methionine ratio coefficient（0.47） and amino acid ratio coefficient
score（75.28） compared to those of S0. In vitro antioxidant tests verified that the selenium protein extract of S3 exhibited strong scavenging capacities against 1，1⁃diphenyl⁃2⁃picrylhydrazyl（DPPH）radicals and hydroxyl radicals，accompanied by favorable ferric⁃reducing power. In conclusion，selenium proteins from high⁃selenium Lentinula edodes prepared by ultrasound⁃assisted enzymatic extraction possess a proportionally balanced amino acid composition，a high proportion of selenoamino acids and excellent antioxidant activity，and have the potential to serve as a novel fungal⁃derived organic selenium nutritional supplement.

Key words: Selenium protein, Selenium?enriched Lentinula edodes, Ultrasound?assisted enzymatic extraction, Amino acid nutritional evaluation, Selenium speciation, Antioxidant activity

摘要： 为优化富硒香菇硒蛋白超声-酶解提取工艺并系统评价其营养品质与抗氧化活性，以蛋白质提取率、总硒提取率为响应指标，通过单因素试验确定参数区间，再采用Box-Behnken响应面法构建回归模型，优化超声-酶解提取工艺；随后将最优工艺应用于4组总硒含量各异的香菇原料（S0：0.3±0.0 mg/kg；S1：9.8±0.4 mg/kg；S2：20.0±0.9 mg/kg；S3：99.6±1.1 mg/kg），对比分析不同原料硒蛋白提取物的硒形态、氨基酸组成及体外抗氧化能力。工艺优化结果表明，最优条件为碱性纤维素酶添加量0.6%（质量比）、料液比1∶25（g/mL）、pH值9.0、酶解温度48 ℃、超声功率300 W、超声时间54 min，该条件下模型预测蛋白质提取率可达50.02%。硒形态分析表明，S3硒蛋白提取物中硒代氨基酸占总硒比例高达86.6%，硒代半胱氨酸为主要赋存形态；氨基酸营养评价结果显示，S3 硒蛋白提取物必需氨基酸/氨基酸总量（40.32%）、蛋氨酸比值系数（0.47）及氨基酸比值系数分（75.28）均优于S0；体外抗氧化试验证实，S3硒蛋白提取物对1，1-二苯基-2-三硝基苯肼（DPPH）自由基、羟基自由基表现出较强的清除能力，且具有良好的铁离子还原力。综上，经超声-酶解法制备的高硒香菇硒蛋白拥有协调均衡的氨基酸组成、高比例硒代氨基酸与优良抗氧化活性，具备开发为一种真菌来源的新型有机硒营养补充剂的潜力。