Optimization of Ultrasonic-assisted Enzymatic Extraction of ACE Inhibitory Peptides from Cyperus esculentus by Response Surface Method
-
摘要: 本文在单因素实验的基础上用响应面法优化了超声辅助酶提取油莎豆ACE（angiotensin converting enzyme）抑制肽工艺，并通过对血管紧张素转化酶的抑制实验选取了最佳辅助酶。结果表明，底物浓度3%、超声处理20 min、酶解温度45 ℃、加酶量5000 U/g、超声功率180 W、酶解3 h是超声波辅助酶法提取油莎豆ACE抑制肽的最佳工艺条件，最佳辅助酶-碱性蛋白酶，在此条件下ACE抑制率为74.16%。本研究为提取油莎豆ACE抑制肽提取了一定理论依据，为进一步研究油莎豆ACE抑制肽奠定了基础。Abstract: In this paper, based on the single-factor experiment, the response surface method was used to optimize the ultrasonic-assisted enzyme extraction process of Cyperus esculentus ACE inhibitory peptide, and the best auxiliary enzyme was selected through the inhibition experiment of angiotensin converting enzyme. The results showed that the optimal process conditions for the ultrasonic-assisted enzymatic extraction of Cyperus esculentus ACE inhibitory peptides were: Substrate concentration 3%, ultrasonic treatment time 20 min, enzymolysis temperature 45 ℃, enzyme addition 5000 U/g, ultrasonic power 180 W, enzymolysis time 3 h, the best auxiliary enzyme-alkaline protease, under this condition, the ACE inhibition rate was 74.16%. This study would provide a theoretical basis for the extraction of Cyperus esculentus ACE inhibitory peptides, which laid a foundation for further research on Cyperus esculentus ACE inhibitory peptide.
-
Key words:
- Cyperus esculentus /
- ultrasound /
- enzymatic hydrolysis /
- ACE inhibitory peptide /
- response surface
-
图 1 油莎豆蛋白等电点的测定
Figure 1. Determination of the isoelectric point of Cyperus esculentus protein
图 2 底物浓度对油莎豆ACE肽抑制率的影响
Figure 2. Effect of substrate concentration on the inhibition rate of Cyperus esculentus ACE peptide
图 3 超声处理时间对油莎豆ACE肽抑制率的影响
Figure 3. Effect of ultrasonic treatment time on inhibition rate of Cyperus esculentus ACE peptide
图 4 超声功率对油莎豆ACE肽抑制率影响
Figure 4. Effect of ultrasonic power on the inhibition rate of Cyperus esculentus ACE peptide
图 5 酶解温度对油莎豆ACE肽抑制率的影响
Figure 5. Effect of enzymatic hydrolysis temperature on the inhibition rate of Cyperus esculentus ACE peptide
图 6 酶解时间对油莎豆ACE肽抑制率的影响
Figure 6. Effect of enzymatic hydrolysis time on the inhibition rate of Cyperus esculentus ACE peptide
图 7 加酶量对油莎豆ACE肽抑制率的影响
Figure 7. Effect of enzyme addition on the inhibition rate of Cyperus esculentus ACE peptide
图 8 酶解时间与酶解温度的交互作用
Figure 8. Interaction between enzymolysis time and enzymolysis temperature
图 9 超声功率与酶解时间的交互作用
Figure 9. Interaction between ultrasonic power and enzymolysis time
图 10 超声功率与酶解温度的交互作用
Figure 10. Interaction between ultrasonic power and enzymolysis temperature
表 1 ACE 抑制率的测定步骤
Table 1. Determination of ACE inhibition rate
试剂 样品 A1（μL） A2（μL） A3（μL） HCl溶液 0 0 200 HHL溶液 200 200 200 ACE抑制剂 50 0 0 硼酸盐缓冲液 0 50 50 ACE溶液 50 50 50 HCl溶液 200 200 0 
下载: 导出CSV
表 2 响应面分析因素及水平
Table 2. Response surface analysis factors and levels
水平 因素 A酶解时间（h） B酶解温度（℃） C超声功率（W） −1 2 40 120 0 3 45 180 1 4 50 240
下载: 导出CSV
表 3 四种蛋白酶对油莎豆ACE抑制肽的抑制率的影响
Table 3. Influence of four proteases on the inhibition rate of Cyperus esculentus ACE inhibitor peptide
酶 ACE抑制率（%） 胃蛋白酶 62.32±0.53 中性蛋白酶 69.15±0.87 碱性蛋白酶 74.45±0.69 木瓜蛋白酶 67.67±0.48
下载: 导出CSV
表 4 Box-Behnken试验设计及其结果
Table 4. Box-Behnken experimental design and results
实验号 A B C ACE抑制率（%） 1 3.00 40.00 240.00 71.82 2 2.00 40.00 180.00 69.91 3 3.00 45.00 180.00 74.13 4 3.00 50.00 120.00 70.34 5 2.00 45.00 120.00 71.24 6 3.00 40.00 120.00 70.46 7 4.00 50.00 180.00 69.35 8 3.00 45.00 180.00 74.51 9 3.00 45.00 180.00 73.92 10 3.00 45.00 180.00 74.38 11 3.00 50.00 240.00 69.97 12 2.00 45.00 240.00 70.75 13 4.00 40.00 180.00 70.89 14 4.00 45.00 120.00 72.25 15 4.00 45.00 240.00 72.14 16 2.00 50.00 180.00 69.43 17 3.00 45.00 180.00 74.53
下载: 导出CSV
表 5 响应面方差分析结果
Table 5. Response surface analysis of variance results
方差来源 平方和 自由度 均方 F P 显著性 模型 55.51 9 6.17 49.40 < 0.0001 *** A 1.36 1 1.36 10.90 0.0131 * B 1.99 1 1.99 15.94 0.0052 ** C 0.019 1 0.019 0.15 0.7080 AB 0.28 1 0.28 2.25 0.1773 AC 0.036 1 0.036 0.29 0.6075 BC 0.75 1 0.75 5.99 0.0442 * A2 12.54 1 12.54 100.42 < 0.0001 *** B2 30.09 1 30.09 240.97 < 0.0001 *** C2 3.99 1 3.99 31.94 0.0008 *** 残差 0.87 7 0.12 失拟项 0.60 3 0.20 2.88 0.1665 不显著 纯误差 0.28 4 0.069 总和 56.39 16 注：***差异高度显著（P<0.001）；**差异极显著（P<0.01）；*差异显著（P<0.05）。
下载: 导出CSV
-
[1] 于红, 敬思群. 油莎豆化学成分及应用研究进展[J]. 食品工业,2015,36(6):242−245. [2] 阳振乐. 油莎豆的特性及其研究进展[J]. 北方园艺,2017,17(392):199−208. [3] Rahul V Manek, Philip F Builders, William M Kolling, et al. Physicochemical and binder properties of starch obtained from Cyperus esculentus[J]. AAPS Pharm Sci Tech,2012,13(2):379−388. doi: 10.1208/s12249-012-9761-z [4] Moonjung Kim, Siwon No, Suk Hoo Yoon. Stereospecific analysis of fatty acid composition of Chufa (Cyperus esculentus L.) tuber oil[J]. Journal of the American Oil Chemists' Society,2007,84(11):1079−1080. doi: 10.1007/s11746-007-1131-8 [5] 陈星, 陈滴, 刘蕾. 油莎豆全成分分析[J]. 食品科技,2009,34(3):165−168. [6] 杨帆, 朱文学. 油莎豆研究现状及展望[J]. 粮食与油脂,2020,33(7):4−6. [7] Hankins C N, Shannon L M. Physical and enzymatic properties of a phytohemagglutinin from mung beans[J]. Journal of Biological Chemistry,1978,253(21):7791−7797. doi: 10.1016/S0021-9258(17)34439-3 [8] Wei-Liang W, Guo-Jie W, Dao-Shuang L, et al. The physiological function and research progress of angiotensin-i-converting enzyme inhibitory petides[J]. Modern Food Science and Technology,2006,22(3):251−254. [9] 孙宁玲. 高血压领域的热点及思考[J]. 中华高血压杂志,2015,23(3):203−205. [10] 罗鹏. 葵花籽ACE抑制肽的分离纯化、结构分析与稳态化研究[D]. 武汉: 华中农业大学, 2018. [11] Martin M, Deussen A. Effects of natural peptides from food proteins on angiotensin converting enzyme activity and hypertension[J]. Critical Reviews in Food Science and Nutrition,2019,59(8):1264−1283. doi: 10.1080/10408398.2017.1402750 [12] Fagyas M, Úri K, Siket I M, et al. New perspectives in the renin-angiotensin-aldosterone system (RAAS) I: Endogenous angiotensin converting enzyme (ACE) inhibition[J]. PLoS One, 2014, 9(4): e87843. [13] Ceren D D, Aysun Y, Funda K G, et al. Angiotensin-i-converting enzyme (ACE)-inhibitory peptides from plants[J]. Nutrients,2017,9(4):316. doi: 10.3390/nu9040316 [14] Wei L W, Guo J W, Dao S L, et al. The physiological function and research progress of angiotensin-i-converting enzyme inhibitory petides[J]. Modern Food Science and Technology,2006(3):251−254. [15] 韩飞, 于婷婷, 周孟良, 等. 酶法生产大豆蛋ACE抑制肽的研究[J]. 食品科学,2008,29(11):369−374. doi: 10.3321/j.issn:1002-6630.2008.11.084 [16] 胡炜东, 蔡永敏, 鲁富宽. 响应面法优化油莎豆粕蛋白抗氧化肽制备工艺[J]. 食品工业,2014,35(2):105−108. [17] 胡炜东, 蔡永敏, 鲁富宽, 等. 响应面分析法优化油莎豆粕蛋白提取工艺[J]. 食品科技,2013,38(6):171−175, 184. [18] Cushman D W, Cheung H S. Spectrophotometric assay and properties of the angiotensin-converting enzyme of rabbit lung[J]. Elsevier,1971,20(7):1637−1648. [19] 杨叶波, 蔡培培, 何文森. 大豆蛋白质的提取技术的研究进展[J]. 广州化工,2015,43(9):26−27. doi: 10.3969/j.issn.1001-9677.2015.09.011 [20] 安传相. 核桃源蛋白ACE抑制肽制备及分离纯化的研究[D]. 贵阳: 贵州大学, 2018. [21] 胡庆娟, 吴光杰, 牛庆川, 等. 响应面试验优化木瓜蛋白酶法脱马齿苋多糖蛋白工艺[J]. 食品科学,2018,39(20):246−252. doi: 10.7506/spkx1002-6630-201820036 [22] 周洁静, 侯银臣, 刘旺旺, 等. 羊胎盘提取残余物免疫肽制备工艺的优化[J]. 食品与发酵工业,2015,41(3):129−134. [23] 韩扬. 超声辅助酶法制备燕麦ACE抑制肽的研究[D]. 北京: 北京工商大学, 2010. [24] 麻成金, 黄伟, 黄群, 等. 复合酶法提取仿栗籽蛋白的工艺优化[J]. 食品科学,2012,33(20):27−32. -
点击查看大图
计量
- 文章访问数: 194
- HTML全文浏览量: 59
- PDF下载量: 10
- 被引次数: 0
