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中国精品科技期刊2020
马二兰,林莹,涂连,等. 芋头球蛋白的提取纯化及其对α-淀粉酶和α-葡萄糖苷酶抑制活性研究[J]. 食品工业科技,2021,42(14):25−32. doi: 10.13386/j.issn1002-0306.2020100266.
引用本文: 马二兰,林莹,涂连,等. 芋头球蛋白的提取纯化及其对α-淀粉酶和α-葡萄糖苷酶抑制活性研究[J]. 食品工业科技,2021,42(14):25−32. doi: 10.13386/j.issn1002-0306.2020100266.
MA Erlan, LIN Ying, TU Lian, et al. Extraction, Purification of Taro Globulin and Its Inhibitory Activity on α-Amylase and α-Glucosidase[J]. Science and Technology of Food Industry, 2021, 42(14): 25−32. (in Chinese with English abstract). doi: 10.13386/j.issn1002-0306.2020100266.
Citation: MA Erlan, LIN Ying, TU Lian, et al. Extraction, Purification of Taro Globulin and Its Inhibitory Activity on α-Amylase and α-Glucosidase[J]. Science and Technology of Food Industry, 2021, 42(14): 25−32. (in Chinese with English abstract). doi: 10.13386/j.issn1002-0306.2020100266.

芋头球蛋白的提取纯化及其对α-淀粉酶和α-葡萄糖苷酶抑制活性研究

Extraction, Purification of Taro Globulin and Its Inhibitory Activity on α-Amylase and α-Glucosidase

  • 摘要: 目的:探究芋头球蛋白体外调节血糖活性。方法:以磷酸缓冲液提取芋头球蛋白,以蛋白质提取率为指标考察了料液比、温度、时间和次数对蛋白质提取的影响,在此基础上采用响应面优化提取工艺。采用DEAE-52离子纤维素柱层析法纯化粗蛋白,通过高效液相色谱法检测纯化所得球蛋白的纯度,并测定其等电点和分子量。研究了芋头球蛋白对α-淀粉酶和α-葡萄糖苷酶的抑制活性及抑制动力学,以阿卡波糖为阳性对照,评价其体外调节血糖活性。结果:最佳提取条件为:料液比1:16 g/mL、温度41 ℃、时间124 min,此时提取率为36.75%±0.31%,得率0.70%±0.04%,纯度85.72%±0.47%。纯化后的球蛋白经高效液相色谱检测得一主峰,纯度为93.27%,得率为0.20%±0.01%,等电点pI=5.6,分子量为22 kDa左右。芋头球蛋白对两种酶的抑制活性与蛋白浓度存在量效关系,对α-淀粉酶的IC50为0.75±0.10 mg/mL,对α-葡萄糖苷酶的IC50为(2.09±0.19) mg/mL,而阿卡波糖对α-淀粉酶的IC50为(0.61±0.13) mg/mL,对α-葡萄糖苷酶的IC50为(0.69±0.16) mg/mL,说明芋头球蛋白对α-淀粉酶略低于阿卡波糖,而对α-葡萄糖苷酶抑制活性远低于阿卡波糖,二者的抑制类型均为可逆性的非竞争性抑制,对α-淀粉酶抑制的Ki=(0.61±0.05) mg/mL,对α-葡萄糖苷酶抑制的Ki=(0.26±0.02) mmol/L。结论:本研究优化了芋头球蛋白的提取工艺,纯化得到芋头球蛋白,研究发现其有一定的体外调节血糖的活性,对功能食品的研发和芋头产品提高附加值具有一定的指导意义。

     

    Abstract: Objective: Exploring taro globulin regulating activity in vitro. Methods: Taro globulin was extracted by phosphate buffer. The effects of solid-liquid ratio, temperature, time and times on protein extraction were investigated with protein extraction rate as an index. Based on the test, response surface methodology was used to optimize the extraction process. The crude protein was purified by DEAE-52 ion cellulose column chromatography. The purity of purified globulin was detected by high performance liquid chromatography, and its isoelectric point and molecular weight were determined. The inhibitory activity and inhibition kinetics of taro globulin on α -amylase and α-glucosidase were studied, and acarbose was used as positive control to evaluate globulin activity of regulating blood glucose in vitro. Results: The optimum extraction conditions were as follows: The solid-to-liquid ratio was 1:16 g/mL, the temperature was 41 ℃, and the time was 124 min. Under these condition, the extraction rate was 36.75%±0.31%, the yield was 0.70%±0.04%, and the purity of the protein was 85.72%±0.47%. The purified globulin was detected by high performance liquid chromatography with a main peak with purity of 93.27%, yield of 0.20%±0.01%, isoelectric point of 5.6 and molecular weight of about 22 kDa. The taro globulin exhibited inhibition on the two enzymes with dose-effect relationship. The IC50 was (0.75±0.10) mg/mL for α-amylase and was (2.09±0.19) mg/mL for α-glucosidase The IC50 of acarbose were (0.61±0.13) mg/mL and (0.69±0.16) mg/mL. The results showed that the inhibitory activity of taro globulin on α-amylase was slightly lower than acarbose, while the inhibitory activity on α-glucosidase was much lower than acarbose. The inhibition type was reversible and uncompetitive inhibitor, with Ki=(0.61±0.05) mg/ml for α-amylase and Ki=(0.26±0.02) mmol/L/L for α-glucosidase. Conclusion: The extraction process of taro globulin was optimized, and taro globulin was purified. It was found that taro globulin has certain activity of regulating blood sugar in vitro, which would have certain guiding significance for the research and development of functional food and the increasing of added value of taro products.

     

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