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中华鳖酶解产物的理化性质及促成骨细胞增殖活性分析

林泽鑫 杨美莲 吴超 洪华贵 杜明

林泽鑫,杨美莲,吴超,等. 中华鳖酶解产物的理化性质及促成骨细胞增殖活性分析[J]. 食品工业科技,2022,43(15):81−88. doi:  10.13386/j.issn1002-0306.2021100235
引用本文: 林泽鑫,杨美莲,吴超,等. 中华鳖酶解产物的理化性质及促成骨细胞增殖活性分析[J]. 食品工业科技,2022,43(15):81−88. doi:  10.13386/j.issn1002-0306.2021100235
LIN Zexin, YANG Meilian, WU Chao, et al. Analysis of the Physicochemical Properties and Osteocyte Proliferative Activity of the Enzymatic Hydrolysis Products of Chinese Soft-shelled Turtle[J]. Science and Technology of Food Industry, 2022, 43(15): 81−88. (in Chinese with English abstract). doi:  10.13386/j.issn1002-0306.2021100235
Citation: LIN Zexin, YANG Meilian, WU Chao, et al. Analysis of the Physicochemical Properties and Osteocyte Proliferative Activity of the Enzymatic Hydrolysis Products of Chinese Soft-shelled Turtle[J]. Science and Technology of Food Industry, 2022, 43(15): 81−88. (in Chinese with English abstract). doi:  10.13386/j.issn1002-0306.2021100235

中华鳖酶解产物的理化性质及促成骨细胞增殖活性分析

doi: 10.13386/j.issn1002-0306.2021100235
基金项目: 兴辽英才计划科技创新领军人才项目(XLYC1802047)。
详细信息
    作者简介:

    林泽鑫(1999−),男,本科,研究方向:蛋白质资源开发与利用,E-mail:lzx2468@163.com

    通讯作者:

    杜明(1977−),男,博士,教授,研究方向:蛋白质资源开发与利用,E-mail:duming@dlpu.edu.cn

  • 中图分类号: TS254.1

Analysis of the Physicochemical Properties and Osteocyte Proliferative Activity of the Enzymatic Hydrolysis Products of Chinese Soft-shelled Turtle

  • 摘要: 本文使用碱性蛋白酶、胃蛋白酶和胰蛋白酶水解中华鳖骨和肉、壳和裙边制备中华鳖功能性肽,并对其主要构象变化、粒度等理化性质进行分析。同时通过测定各酶解物对MC3T3-E1细胞增殖情况的影响对其潜在的骨密度调节活性进行评价。结果显示,中华鳖壳和裙边干粉蛋白含量为54.18%,骨和肉干粉蛋白质含量为80.38%。酶解2 h后,碱性蛋白酶和胰蛋白酶的酶解产物分子量集中分布于1000 Da以下,胃蛋白酶酶解产物分子量分布较为均匀。酶解产物的结构以无规则卷曲为主,粒径集中分布于20和200 nm区域,说明酶解较为充分。酶解产物中赖氨酸、缬氨酸、苯丙氨酸、亮氨酸和苏氨酸等必需氨基酸的含量较高,非必需氨基酸中甘氨酸、丙氨酸和脯氨酸的含量相对较高。其中,胃蛋白酶酶解中华鳖骨和肉后必需氨基酸的总量达39.81%。细胞实验结果显示,六种酶解产物均有一定的促成骨细胞增殖活性,壳和裙边酶解物的增殖活性强于骨肉酶解物,其中胃蛋白酶酶解的壳和裙边产物活性最强,说明中华鳖酶解产物具有潜在的骨密度调节活性。本研究对于中华鳖等产品的深加工提供了可行性分析和理论参考。
  • 图  1  碱性蛋白酶酶解中华鳖不同部位过程中pH随时间的变化情况

    Figure  1.  Changes in pH with time during enzymatic hydrolysis of different parts of the Chinese soft-shelled turtle by alkaline protease

    注:A:碱性蛋白酶酶解骨和肉;B:碱性蛋白酶酶解壳和裙边。

    图  2  胃蛋白酶酶解中华鳖不同部位过程中pH随时间的变化情况

    Figure  2.  Changes in pH with time during enzymatic hydrolysis of different parts of the Chinese soft-shelled turtle by pepsin

    注:A:胃蛋白酶酶解骨和肉;B:胃蛋白酶酶解壳和裙边。

    图  3  胰蛋白酶酶解中华鳖不同部位过程中pH随时间的变化情况

    Figure  3.  Changes in pH with time during enzymatic hydrolysis of different parts of the Chinese soft-shelled turtle by trypsin

    注:A:胰蛋白酶酶解骨和肉;B:胰蛋白酶酶解壳和裙边。

    图  4  酶解中华鳖不同部位酶解物的分子量分布

    Figure  4.  The molecular weight distribution of the hydrolysates from different parts of the Chinese soft-shelled turtle

    图  5  圆二色光谱分析结果

    Figure  5.  Circular dichroic spectrum analysis results

    图  6  内源荧光光谱分析结果

    Figure  6.  Analysis results of endogenous fluorescence spectroscopy

    图  7  动态光散射分析结果

    Figure  7.  Dynamic light scattering analysis results

    图  8  24、48、72 h后三种浓度下不同酶解产物成骨细胞存活率

    Figure  8.  The survival rate of osteoblasts from different enzymatic hydrolysis products at three concentrations after 24, 48, 72 h

    注:A:24 h;B:48 h;C:72 h;Control:空白对照组;*代表与空白对照组相比,显著差异,P<0.05。

    表  1  酶的最适pH和温度

    Table  1.   Optimal pH and temperature for enzymes

    名称pH最适温度(℃)
    碱性蛋白酶10.050
    胃蛋白酶2.037
    胰蛋白酶8.545
    下载: 导出CSV

    表  2  中华鳖蛋白酶解物的总氨基酸含量

    Table  2.   Total amino acid content of enzymatic hydrolysate of Chinese soft-shelled turtle protein

    氨基酸种类氨基酸含量(mg/g酶解物)
    必需氨基酸非必需氨基酸壳和裙边骨和肉
    胰蛋白酶胃蛋白酶碱性蛋白酶胰蛋白酶胃蛋白酶碱性蛋白酶
    苏氨酸26.493.6836.1834.2131.7626.10
    半胱氨酸0.000.000.000.0085.790.00
    赖氨酸25.614.3837.7345.5257.2124.90
    蛋氨酸0.000.002.142.650.004.40
    苯丙氨酸34.719.7842.6440.4154.3528.76
    异亮氨酸9.761.0015.6115.6613.819.53
    亮氨酸20.732.1633.4830.9227.9319.40
    组氨酸10.531.6215.1516.7221.739.37
    色氨酸
    酪氨酸28.3110.7849.0247.6554.7219.36
    缬氨酸30.066.8932.5530.8128.9830.45
    丝氨酸42.316.2245.2039.7344.6642.39
    精氨酸48.787.9940.8057.4961.4973.75
    天冬氨酸67.8511.9890.8878.1298.0061.37
    甘氨酸142.8223.61103.1790.06130.70156.78
    谷氨酸102.1417.52146.88127.03145.7098.51
    丙氨酸95.2215.7888.9478.630.0097.61
    脯氨酸96.5316.1175.3061.1588.2598.81
    必需氨基酸总值186.2040.30264.51264.55376.29172.27
    非必需氨基酸总值595.6699.22591.15532.22568.81629.22
    注:“—”表示未检出,表3同。
    下载: 导出CSV

    表  3  中华鳖蛋白的酶解物游离氨基酸含量

    Table  3.   Free amino acid content of enzymatic hydrolysate of Chinese soft-shelled turtle protein

    氨基酸种类氨基酸含量(mg/g酶解物)
    必需氨基酸非必需氨基酸壳和裙边骨和肉
    胰蛋白酶胃蛋白酶碱性蛋白酶胰蛋白酶胃蛋白酶碱性蛋白酶
    苏氨酸0.000.150.000.000.260.44
    半胱氨酸0.180.160.240.000.300.88
    赖氨酸0.900.071.631.351.692.03
    蛋氨酸0.420.000.000.590.000.82
    苯丙氨酸5.251.8010.315.838.0510.18
    异亮氨酸0.000.000.000.000.000.00
    亮氨酸0.030.000.030.000.000.00
    组氨酸0.000.021.850.000.010.92
    色氨酸
    酪氨酸1.220.111.851.141.6923.49
    缬氨酸1.080.190.930.580.370.63
    丝氨酸0.460.160.000.570.190.00
    精氨酸0.000.000.000.460.000.08
    天冬氨酸1.010.170.581.260.410.74
    甘氨酸0.020.010.000.040.060.01
    谷氨酸1.210.270.811.170.390.62
    丙氨酸0.000.000.000.100.000.00
    脯氨酸0.000.000.000.000.000.00
    总游离氨基酸11.783.1318.2213.0713.4040.83
    苦味氨基酸6.782.0213.126.998.4312.55
    香味氨基酸2.210.441.392.420.791.36
    甜味氨基酸0.490.320.000.710.500.45
    下载: 导出CSV
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出版历程
  • 收稿日期:  2021-10-22
  • 网络出版日期:  2022-06-17
  • 刊出日期:  2022-08-03

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