萌发过程大豆蛋白质动态变化及营养价值的研究
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摘要
大豆(Glycine max(L.)Merr)系豆科大豆属栽培种,栽培历史悠久,是蛋白质的良好来源。但是,蛋氨酸的缺乏影响着大豆蛋白的利用,胰蛋白酶抑制剂的存在能够减少蛋白质的利用并降低大豆食品的营养效用。因此,降低大豆胰蛋白酶抑制剂的活性、平衡大豆蛋白质的氨基酸组成对提高大豆蛋白质的利用率至关重要。研究证实,萌发可提高大豆营养价值,降低胰蛋白酶抑制剂的活力。因此,探索高利用率、高营养效用大豆蛋白的适宜萌发条件和萌发阶段至关重要。
本研究以此为立足点,以中农056444品系大豆为材料,研究大豆种子萌发过程中吸水规律、蛋白质代谢、胰蛋白酶抑制剂变化,通过建立以可溶性蛋白质含量和胰蛋白酶抑制剂比活力的回归模型来探寻适宜的萌发条件,对优化条件处理的大豆种子进行深入分析,研究其营养价值,找到营养价值最佳的萌发条件,结果如下:
1)大豆种子萌发过程吸水阶段的转折点分别在吸胀12 h和吸水27 h;萌发适宜pH为7.5;适宜的萌发方式是半浸种方式;适宜萌发温度是20℃。
2)萌发过程中大豆可溶性蛋白质的含量均高于风干种子的含量,峰值分别出现在萌发12 h和72 h,增幅分别达到16%和20%;蛋白酶和肽酶活性在萌发0～39 h之间变化平稳,39～72 h之间急剧上升,72 h时蛋白酶活力较风干种子增加了65%,肽酶活力增加了78%,随后活性均下降;肽和氨态氮在整个萌发过程中呈上升趋势,萌发96 h时肽含量较风干种子增加了39%,氨态氮含量增加了152%。pH7.5、萌发24 h条件下萌发温度为20℃时可溶性蛋白含量最低,最高值(30℃)比其增幅为8.3%。
可溶性蛋白质峰值出现在pH 6.0和pH 8.5,最高值与最低值之间差异为5.1%。内肽酶最适pH值为pH 5.0～6.0,较最低值高出11.7%,外肽酶最适pH值为pH 9.0,较最低值高出39%;pH 7.0和9.0时肽含量最大,氨态氮pH 6.0时达最大值。
3)萌发对胰蛋白酶抑制剂比活力的影响大于对胰蛋白酶抑制剂活力的影响,萌发0～72 h之间胰蛋白酶抑制剂活性变化不大,萌发72 h时胰蛋白酶抑制剂活力降至风干种子的90%,比活力降至风干种子的68%;萌发至96 h时胰蛋白酶抑制剂活力下降至风干种子的69%,比活力下降至风干种子的53%。萌发温度和浸种液pH对胰蛋白酶抑制剂的影响没有萌发时间大,最大差异不超过10%。
4)内肽酶活性在萌发9 h时较风干种子提高了24%,萌发9 h之后内肽酶活性变化不大,但整个萌发过程中均处于高水平状态;处理范围内最适萌发温度和最适浸种液pH条件下的内肽酶活性也比最不适的萌发温度和浸种液pH条件下的内肽酶活性提高不超过6%。
5)回归模型所描述的可溶性蛋白质、胰蛋白酶抑制剂比活力随萌发温度、时间和pH的变化规律是有效和显著的,能够可靠地表征可溶性蛋白质和胰蛋白酶抑制剂比活力对萌发温度、时间和pH的变化规律。
6)萌发大豆较风干种子的必需氨基酸/总氨基酸最高增幅为5%,必需氨基酸/非必需氨基酸的值最高增幅为15%,必需氨基酸模式更加符合FAO推荐模式,AAS、CS、EAAI值均不同程度增加增加,结合蛋白质功效比,经过综合比较,65 h,27℃、pH 7.5处理的营养价值最优,其次是20℃、12 h、pH 7.5条件处理。
Soybean(Glycine max(L.) Merr) being as a soja subgenus cultivar which belongs to pulse family has a long time and wildcultivation in the world,and it is a good food providing various proteins. Wherease the scare of methionine affects the utilation rate of soybean nutition,among which,trypsin inhibitor,as a digeative enzyme during development,is an important anti-nutrition factor.Trypsin protease inhibitor can decrease the utilization of protein,and result in the loss of soybean nutrition for human beings.So decreasing the trypsin inhibitor proteinase activity and blancing the composition of amino acid is necessary and important to improve usage rate.The study showes that germination of seed can enhance the nutrition utilization derive from inhibition of the activation of trypsin protease inhibitor. Given the health of human beings,it is necessary to find optimal germination conditions which absolutely improve soybean protein nutrition in the future.
Considered the importance of this study,ZhongNong 056444 was used as a material to study the tendency of imbibiton,metabolism of proteins,the change of trypsin protease inhibitor and peptidase.A secondary orthogonal regression design of three factors model was constructed based on the data of soluble proteins and species activity of trypsin protease inhibitor to explore the optimal conditions of germination.The result of this study was shown as following:
1) The germination experiment and protein metabolism result was that imbibition processes could be classified into three phases.The reversal points appeared at 12 h and 27 h respectively during germination,and the optimal conditions was Ph 7.5 and 20℃under semi-seed soaking condition.
2) Temperature,pH and germination time affected the soybean protein metabolism during germination,and among which,the time and temperature were key factors.The maximal soluble protein appeared in 12 h and 72 h in imbited seed,and it was enhanced 16%and 20%respectively than that of dry seed,The activity of protease and peptidase showed no significant change when seed was imbibed between 0～39 h,however,novel enhancement was observed between 39～72 h,and decreased in the later time.During this period,proteinase and content in 72 h was improved 65%and 78%respectively during imbibition than that of dry seed.Peptide and N-anmino enhanced their level in 96 h in germinated seed,and enhanced their level for 39%and 152 respectively than dry seed.The soluble protein content was minimalest when seed was geriminated under 20℃for 24 h.
In addition,the maximum of soluble protein content was shown in pH 6.0 and pH 8.5,and was improved for a 5.1%.The most optimal pH condition for peptidase was 5.0～6.0,and 11.7% improvement was observed,pH 9.0 is the optimal condition for the activity of exopeptidases with 39% enhancement.The content of peptide and N-amino acid was the most in pH 6.0 than that of others.
3) Significant effect on the activity or species activity of trypsin protease was found when the seed was germinated,and the species activity was the most sensitivity in the germinated seed.No change was found at 0～72 h.However,the activity of trypsin protease inhibitor began to decrease after 72 h imbibition,.The activity and species activity of trypsin protease inhibitor was decreased to a 90%and 68%respectively in this time than the dry seed,and was only 69%and 53%after 96 h imbibtion.No significant effect was found for the pH on the activity of trypsin protease with no more than 10% enhancement.
4) From the result of regression equation,the trend of soluble protein and trypsin protease activity under treatment of temperature,pH and germinated time was notable,and it could correctly elucidate the tendency of soluble protein and the species activity of trypsin protease inhibitor during germination under above conditions.
5) The increase of endopeptidases was enhanced for a 24%at 9 h,while no change was showed in the following time,although high level was showed during experiment time.Under optimal temperature and pH conditions,the activity of endopeptidases was only improved no more than 6%than unavailable conditions.
6) The 5%improvement of essential amino acid content/total amino acid amino was observed in the germinated seeds.The model of essential amino acid was corresponding to the FAO recommend model, and the AAS,CS and EAAI showen increase at different level.Combined with efficiency ratio,the protein nutrition of soybean would be the most great under the 65 h,27℃and pH 7.5.Together,the data of this study probably provided the new information to improve the health of human beings.
本研究以此为立足点,以中农056444品系大豆为材料,研究大豆种子萌发过程中吸水规律、蛋白质代谢、胰蛋白酶抑制剂变化,通过建立以可溶性蛋白质含量和胰蛋白酶抑制剂比活力的回归模型来探寻适宜的萌发条件,对优化条件处理的大豆种子进行深入分析,研究其营养价值,找到营养价值最佳的萌发条件,结果如下:
1)大豆种子萌发过程吸水阶段的转折点分别在吸胀12 h和吸水27 h;萌发适宜pH为7.5;适宜的萌发方式是半浸种方式;适宜萌发温度是20℃。
2)萌发过程中大豆可溶性蛋白质的含量均高于风干种子的含量,峰值分别出现在萌发12 h和72 h,增幅分别达到16%和20%;蛋白酶和肽酶活性在萌发0～39 h之间变化平稳,39～72 h之间急剧上升,72 h时蛋白酶活力较风干种子增加了65%,肽酶活力增加了78%,随后活性均下降;肽和氨态氮在整个萌发过程中呈上升趋势,萌发96 h时肽含量较风干种子增加了39%,氨态氮含量增加了152%。pH7.5、萌发24 h条件下萌发温度为20℃时可溶性蛋白含量最低,最高值(30℃)比其增幅为8.3%。
可溶性蛋白质峰值出现在pH 6.0和pH 8.5,最高值与最低值之间差异为5.1%。内肽酶最适pH值为pH 5.0～6.0,较最低值高出11.7%,外肽酶最适pH值为pH 9.0,较最低值高出39%;pH 7.0和9.0时肽含量最大,氨态氮pH 6.0时达最大值。
3)萌发对胰蛋白酶抑制剂比活力的影响大于对胰蛋白酶抑制剂活力的影响,萌发0～72 h之间胰蛋白酶抑制剂活性变化不大,萌发72 h时胰蛋白酶抑制剂活力降至风干种子的90%,比活力降至风干种子的68%;萌发至96 h时胰蛋白酶抑制剂活力下降至风干种子的69%,比活力下降至风干种子的53%。萌发温度和浸种液pH对胰蛋白酶抑制剂的影响没有萌发时间大,最大差异不超过10%。
4)内肽酶活性在萌发9 h时较风干种子提高了24%,萌发9 h之后内肽酶活性变化不大,但整个萌发过程中均处于高水平状态;处理范围内最适萌发温度和最适浸种液pH条件下的内肽酶活性也比最不适的萌发温度和浸种液pH条件下的内肽酶活性提高不超过6%。
5)回归模型所描述的可溶性蛋白质、胰蛋白酶抑制剂比活力随萌发温度、时间和pH的变化规律是有效和显著的,能够可靠地表征可溶性蛋白质和胰蛋白酶抑制剂比活力对萌发温度、时间和pH的变化规律。
6)萌发大豆较风干种子的必需氨基酸/总氨基酸最高增幅为5%,必需氨基酸/非必需氨基酸的值最高增幅为15%,必需氨基酸模式更加符合FAO推荐模式,AAS、CS、EAAI值均不同程度增加增加,结合蛋白质功效比,经过综合比较,65 h,27℃、pH 7.5处理的营养价值最优,其次是20℃、12 h、pH 7.5条件处理。
Soybean(Glycine max(L.) Merr) being as a soja subgenus cultivar which belongs to pulse family has a long time and wildcultivation in the world,and it is a good food providing various proteins. Wherease the scare of methionine affects the utilation rate of soybean nutition,among which,trypsin inhibitor,as a digeative enzyme during development,is an important anti-nutrition factor.Trypsin protease inhibitor can decrease the utilization of protein,and result in the loss of soybean nutrition for human beings.So decreasing the trypsin inhibitor proteinase activity and blancing the composition of amino acid is necessary and important to improve usage rate.The study showes that germination of seed can enhance the nutrition utilization derive from inhibition of the activation of trypsin protease inhibitor. Given the health of human beings,it is necessary to find optimal germination conditions which absolutely improve soybean protein nutrition in the future.
Considered the importance of this study,ZhongNong 056444 was used as a material to study the tendency of imbibiton,metabolism of proteins,the change of trypsin protease inhibitor and peptidase.A secondary orthogonal regression design of three factors model was constructed based on the data of soluble proteins and species activity of trypsin protease inhibitor to explore the optimal conditions of germination.The result of this study was shown as following:
1) The germination experiment and protein metabolism result was that imbibition processes could be classified into three phases.The reversal points appeared at 12 h and 27 h respectively during germination,and the optimal conditions was Ph 7.5 and 20℃under semi-seed soaking condition.
2) Temperature,pH and germination time affected the soybean protein metabolism during germination,and among which,the time and temperature were key factors.The maximal soluble protein appeared in 12 h and 72 h in imbited seed,and it was enhanced 16%and 20%respectively than that of dry seed,The activity of protease and peptidase showed no significant change when seed was imbibed between 0～39 h,however,novel enhancement was observed between 39～72 h,and decreased in the later time.During this period,proteinase and content in 72 h was improved 65%and 78%respectively during imbibition than that of dry seed.Peptide and N-anmino enhanced their level in 96 h in germinated seed,and enhanced their level for 39%and 152 respectively than dry seed.The soluble protein content was minimalest when seed was geriminated under 20℃for 24 h.
In addition,the maximum of soluble protein content was shown in pH 6.0 and pH 8.5,and was improved for a 5.1%.The most optimal pH condition for peptidase was 5.0～6.0,and 11.7% improvement was observed,pH 9.0 is the optimal condition for the activity of exopeptidases with 39% enhancement.The content of peptide and N-amino acid was the most in pH 6.0 than that of others.
3) Significant effect on the activity or species activity of trypsin protease was found when the seed was germinated,and the species activity was the most sensitivity in the germinated seed.No change was found at 0～72 h.However,the activity of trypsin protease inhibitor began to decrease after 72 h imbibition,.The activity and species activity of trypsin protease inhibitor was decreased to a 90%and 68%respectively in this time than the dry seed,and was only 69%and 53%after 96 h imbibtion.No significant effect was found for the pH on the activity of trypsin protease with no more than 10% enhancement.
4) From the result of regression equation,the trend of soluble protein and trypsin protease activity under treatment of temperature,pH and germinated time was notable,and it could correctly elucidate the tendency of soluble protein and the species activity of trypsin protease inhibitor during germination under above conditions.
5) The increase of endopeptidases was enhanced for a 24%at 9 h,while no change was showed in the following time,although high level was showed during experiment time.Under optimal temperature and pH conditions,the activity of endopeptidases was only improved no more than 6%than unavailable conditions.
6) The 5%improvement of essential amino acid content/total amino acid amino was observed in the germinated seeds.The model of essential amino acid was corresponding to the FAO recommend model, and the AAS,CS and EAAI showen increase at different level.Combined with efficiency ratio,the protein nutrition of soybean would be the most great under the 65 h,27℃and pH 7.5.Together,the data of this study probably provided the new information to improve the health of human beings.
引文
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