黑豆蛋白的分级提取及黑豆花色苷的成分鉴定

Fractional Extraction of Protein and Characterization of Anthocyanins from Black Soybean Seeds

以黑豆为原料,添加不同浓度(0、5、10、15、20、25 mmol/L)的不同二价阳离子(Ca^(2+)、Mg^(2+)和Zn^(2+))分级提取黑豆蛋白,经冷冻干燥后用凯氏定氮法测得蛋白质量浓度,并对所得蛋白级分(7S、11S)的纯度和得率进行分析。结果表明,黑豆提取过程中所加入二价阳离子的类型及浓度显著影响7S和11S球蛋白的得率和纯度。利用综合平衡法,最终选定添加10 mmol/L的Mg2+分级提取黑豆球蛋白,得到7S蛋白纯度和得率分别为(86.29±3.25)%和(2.90±0.14)%,11S蛋白纯度和得率分别为(87.42±3.30)%和(9.11±0.28)%。用乙醇法提取黑豆花色苷,测得黑豆中总花色苷含量为(0.58±0.03)mg/g,总酚含量为(2.22±0.12)mg/g,对黑豆花色苷进行高效液相色谱-串联质谱分析,其组成成分为飞燕草色素-3-O-葡萄糖苷、矢车菊色素-3-O-葡萄糖苷、牵牛花色素-3-O-葡萄糖苷和锦葵花色素-3-O-葡萄糖苷。

Crude protein extract from black soybean seeds was fractionated by addition of different concentrations(0,5,10,15,20and25mmol/L)of divalent cations(Ca2+,Mg2+and Zn2+).After the precipitated protein fractions(7S globulinand11S globulin)were freeze dried,total nitrogen was determined by the Kjeldahl method and the yield and purity ofthe two fractions were measured.The results revealed that the type and concentration of divalent cations significantlyaffected the yield and purity of7S and11S globulin.Based on comprehensive consideration of yield and purity,10mmol/Lof Mg2+was selected to extract black soybean globulin.The purity and yield of the obtained7S protein were(86.29±3.25)%and(2.90±0.14)%,respectively,while the purity and yield of the obtained11S protein were(87.42±3.30)%and(9.11±0.28)%,respectively.Anthocyanins from black soybean seeds were extracted with ethanol.It turned out that thecontent of anthocyanins was(0.58±0.03)mg/g and total phenol was(2.22±0.12)mg/g.By high performance liquidchromatography tandem-mass spectrometry(HPLC-MS/MS)analysis,the isolated anthocyanins were characterized asdelphinidin-3-O-glucoside,cyanidin3-O-glucoside,petunidin3-O-glucoside and mallow-3-O-glucoside,respectively.