酱油渣异黄酮抗氧化功效成分纯化研究

以酱油渣为原料，采用乙醇提取法提取酱油渣中大豆异黄酮，对提取条件进行了优化。比较了醇沉法、等电点沉淀法、浓缩离心法、正丁醇法、二氯甲烷法、乙酸乙酯萃取法等6种方法对异黄酮粗提物的纯化效果。通过高效液相鉴定了纯化产物的异黄酮单体组成，并采用邻苯三酚氧化法和DPPH法评价了异黄酮纯化物的体外抗氧化活性。结果表明：优化条件下提取的异黄酮粗提物中异黄酮质量分数为1.21%、提取率为（85.36±0.09）%。6种纯化方法中，浓缩离心法所得异黄酮产物中异黄酮纯度为7.75%，去除蛋白质效果最佳；二氯甲烷萃取法所得异黄酮产物中异黄酮纯度为27.74%，去除总糖和灰分效果最佳；两者联用后所得纯化产物的异黄酮纯度为44.88%，其主要由染料木素、大豆黄素和大豆素组成，质量分数分别为67.37%、9.33%、23.30%。在质量浓度为1 mg/mL时，异黄酮纯化物对超氧阴离子、DPPH自由基的清除率分别为67.65%、89.18%，显著高于异黄酮粗提物（P＜0.05），其IC50分别为0.34、0.26 mg/mL，说明异黄酮纯化物具有一定的抗氧化活性，有望作为一种潜在的天然抗氧化剂应用于功能性食品中。

Purification of Isoflavones from Soy Sauce Residue and Characterization of Antioxidant Components

The soybean isoflavone was firstly extract from soy sauce residue by ethanol extraction method, and the extraction conditions were optimized. The crude isoflavone extract was further purified by 6 methods including alcohol precipitation, isoelectric precipitation, concentrated centrifugation, n-butanol, dichloromethane, and ethyl acetate extraction. Finally, the composition of isoflavone monomer after purification was identified by high performance liquid phase, and its in vitro antioxidation was evaluated by the pyrogallol oxidation and DPPH methods. The optimized extraction of crude isoflavone yielded the isoflavone content of 1.21% and the extraction rate of (85.36±0.09)%. Among the 6 purification methods, the isoflavone content obtained by the concentrated centrifugation method was 7.75% with best removal of protein. The isoflavone content obtained by the dichloromethane extraction method was 27.74% with best removal of total sugar and ash. The purity of the isoflavone was 44.88% when the two methods combined, and it was composed by genistein, glycintein and daidzein, accounting for 67.37% and 9.33%, 23.30%, respectively. The scavenging rates of purified isoflavones(1 mg/mL) to superoxide and DPPH free radicals were 67.65% and 89.18%, which were significantly higher than isoflavones crude extracts(P<0.05). And their IC₅₀ were 0.34 and 0.26 mg/mL. The isoflavone purified products achieve certain antioxidant activity, expected to be used as potential natural antioxidants in functional foods.