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中国生物工程杂志

CHINA BIOTECHNOLOGY
中国生物工程杂志
中国生物工程杂志  2022, Vol. 42 Issue (11): 88-98    DOI: 10.13523/j.cb.2209040
生物质资源     
向日葵籽壳黑色素的分离提取及生物活性研究*
李超峰1,2,吴育萍1,李双双1,胥凤梅1,章心怡1,王伟中1,2,唐伯平1,2,**()
1 盐城师范学院湿地学院 盐城 224007
2 盐城师范学院 江苏省滩涂生物资源与环境保护重点实验室 盐城 224007
Isolation and Biological Activity of Melanin from Sunflower Seed Shell
LI Chao-feng1,2,WU Yu-ping1,LI Shuang-shuang1,XU Feng-mei1,ZHANG Xin-yi1,WANG Wei-zhong1,2,TANG Bo-ping1,2,**()
1 School of Wetlands, Yancheng Teachers University, Yancheng 224007, China
2 Jiangsu Key Laboratory of Coastal Wetland Bioresources and Environmental Protection, Yancheng Teachers University, Yancheng 224007, China
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摘要：

目的:天然黑色素安全性高,兼具多种生物活性,深受消费者青睐,发展前景十分广阔。研究向日葵籽壳黑色素制备条件,以及抗氧化和吸附重金属生物活性,为其进一步应用提供理论基础。方法: 采用热碱提取、酸水解、有机溶剂洗涤和反复沉淀法,结合单因素与响应面分析法实现了黑色素的高效制备,利用氯化硝基四氮唑蓝还原法、分光光度法和静态吸附法进行抗氧化活性和吸附活性测定。结果: 成功制备了向日葵籽壳黑色素。提取结果表明最优工艺为氢氧化钠浓度0.11 mol/L、提取温度75.19℃、提取时间181.16 min、料液比为15.77∶1,向日葵籽壳黑色素得率为2.95%,与预测值相近,说明模型拟合良好,该优化工艺准确可行。抗氧化活性测定结果表明,向日葵籽壳黑色素清除超氧阴离子自由基、DPPH自由基的能力和还原力都高于合成型黑色素。吸附活性测定结果表明,向日葵籽壳黑色素对Pb2+、Cu2+和Cr3+的吸附效率高于合成型黑色素。结论: 与其合成型黑色素相比,天然黑色素既可作为着色剂,又兼抗氧化剂和吸附剂,且不亚于合成型黑色素的效果,可用于代替合成型黑色素开发的重要候选。
关键词： 向日葵籽壳;  黑色素;  分离提取;  抗氧化活性;  吸附作用    
Abstract:

Objective: Natural edible pigments with high safety and low toxicity usually possess various nutritional and pharmacological effects, and they have huge practical application value in the market. In this paper, the extraction condition, antioxidant activity, which included DPPH radical-scavenging activity, superoxide radical (O2-)-scavenging activity and reducing power, and metal chelating activity of melanin from sunflower seed shell were investigated in detail to provide theoretical basis for its future application. Methods: In this paper, the efficient preparation of melanin from sunflower seed shell was achieved using hot alkali extraction by single-factor experiments combined with response surface methodology (RSM), acid hydrolysis, washing with organic solvents, re-dissolution and re-precipitation. The antioxidant activity and adsorption activity were determined by nitrotetrazolium nlue chloride reduction method, spectrophotometry and static adsorption. Results: Melanin from sunflower seed shell was successfully prepared. The optimal conditions were determined as follows: NaOH concentration, 0.11 mol/L; liquid-to-solid ratio, 15.77∶1; extraction time, 181.16 min; and extraction temperature, 75.19℃. Under these conditions, the experimental yield of melanin was 2.95%, which was well matched with the value predicted by the developed model. The ability to scavenge DPPH radical, superoxide radical (O2-) and the reducing power of melanin from sunflower seed shell were stronger than those of synthetic melanin. Furthermore, the adsorption efficiency of melanin from sunflower seed shell to Pb2+, Cu2+ and Cr3+ were higher than that of synthetic melanin. Conclusion: Compared with synthetic melanin, natural melanin from sunflower seed shell could be used as colorant, antioxidant and adsorbent, and was more effective. Therefore, melanin from sunflower seed shell was a potential replacement of synthetic melanin.
Key words: Sunflower seed shell    Melanin    Extraction    Antioxidant activity    Absorption
收稿日期: 2022-09-15 出版日期: 2022-12-07
ZTFLH:  Q819  
基金资助: *国家自然科学基金面上项目(31672267);江苏省社会发展重点研发计划(BE2020673);江苏省自然科学基金青年科学基金(BK20150422)
通讯作者: **电子信箱:962664124@qq.com; boptang@163.com   
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李超峰
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章心怡
王伟中
唐伯平

引用本文:

李超峰, 吴育萍, 李双双, 胥凤梅, 章心怡, 王伟中, 唐伯平. 向日葵籽壳黑色素的分离提取及生物活性研究*[J]. 中国生物工程杂志, 2022, 42(11): 88-98.

LI Chao-feng, WU Yu-ping, LI Shuang-shuang, XU Feng-mei, ZHANG Xin-yi, WANG Wei-zhong, TANG Bo-ping. Isolation and Biological Activity of Melanin from Sunflower Seed Shell. China Biotechnology, 2022, 42(11): 88-98.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.2209040    或    https://manu60.magtech.com.cn/biotech/CN/Y2022/V42/I11/88

图1  NaOH浓度对黑色素提取的影响
图2  料液比对黑色素提取的影响
图3  提取温度对黑色素提取的影响
图4  提取时间对黑色素提取的影响
试验号 X1NaOH浓度/(mol/L) X2温度/℃ X3时间/h X4料液比 得率/%
1 0.1 60 2 15∶1 2.18±0.02
2 0.1 75 2 10∶1 2.18±0.03
3 0.1 75 4 20∶1 2.37±0.05
4 0.1 75 3 15∶1 2.98±0.02
5 0.2 60 3 15∶1 2.42±0.03
6 0.1 75 3 15∶1 2.96±0.04
7 0.2 75 3 10∶1 2.32±0.02
8 0.1 90 2 15∶1 2.30±0.03
9 0.0 75 3 10∶1 2.18±0.03
10 0.0 75 2 15∶1 2.26±0.01
11 0.2 75 2 15∶1 2.25±0.02
12 0.2 90 3 15∶1 2.32±0.02
13 0.1 75 2 20∶1 2.35±0.02
14 0.1 60 3 20∶1 2.41±0.03
15 0.1 75 4 10∶1 2.21±0.02
16 0.0 60 3 15∶1 2.09±0.01
17 0.2 75 4 15∶1 2.47±0.01
18 0.1 75 3 15∶1 2.95±0.02
19 0.1 90 3 20∶1 2.41±0.01
20 0.1 75 3 15∶1 2.94±0.02
21 0.2 75 3 20∶1 2.55±0.02
22 0.0 90 3 15∶1 2.32±0.02
23 0.0 75 3 20∶1 2.33±0.01
24 0.1 90 4 15∶1 2.26±0.01
25 0.1 75 3 15∶1 2.97±0.00
26 0.0 75 4 15∶1 2.10±0.03
27 0.1 90 3 10∶1 2.23±0.01
28 0.1 60 4 15∶1 2.26±0.02
29 0.1 60 3 10∶1 2.22±0.02
表1  Box-Behnken试验设计方案与结果
变异来源 平方和 自由度 均方 F值 P值 显著性
模型 2.14 14 0.15 451.00 <0.000 1 **
X1 0.092 1 0.092 271.27 <0.000 1 **
X2 5.633×10-3 1 5.633×10-3 16.63 0.001 1 **
X3 1.875×10-3 1 1.875×10-3 5.54 0.033 8 *
X4 0.097 1 0.097 286.99 <0.000 1 **
X1X2 0.027 1 0.027 80.38 <0.000 1 **
X1X3 0.036 1 0.036 106.59 <0.000 1 **
X1X4 1.600×10-3 1 1.600×10-3 4.72 0.047 4 *
X2X3 3.600×10-3 1 3.600×10-3 10.63 0.005 7 **
X2X4 2.500×10-5 1 2.500×10-3 0.074 0.789 8
X3X4 2.500×10-5 1 2.500×10-3 0.074 0.789 8
X 1 2 0.66 1 0.66 1 961.13 <0.000 1 **
X 2 2 0.77 1 0.77 2 263.04 <0.000 1 **
X 3 2 0.90 1 0.90 2 657.41 <0.000 1 **
X 4 2 0.59 1 0.59 1 738.04 <0.000 1 **
残差 4.742×10-3 14 3.387×10-4
失拟项 3.742×10-3 10 3.742×10-4 1.50 0.371 4
纯误差 1.000×10-3 4 2.500×10-4
总误差 2.14 28
表2  回归模型的方差分析及回归系数的显著性检验
图5  各因素交互作用对黑色素得率影响的响应曲面
序号 分析 结果
1 颜色 黑色
2 水 不溶
3 盐酸溶液 不溶
4
 甲醇、乙醇、氯仿、乙酸乙酯、石油醚、
正丁醇、丙酮等有机溶剂		 不溶
5
 氨水、NaOH、KOH、NaHCO3、
Na2CO3等碱性溶液		 溶解
6 H2O2、NaOCl、KMnO4、K2Cr2O7溶液 脱色
7 FeCl3溶液 棕色絮状物
表3  向日葵籽壳黑色素的理化性质
图6  向日葵籽壳黑色素的紫外-可见光谱
图7  向日葵籽壳黑色素对DPPH自由基的清除效果
图8  向日葵籽壳黑色素对超氧阴离子自由基的清除效果
图9  向日葵籽壳黑色素的还原力
图10  向日葵籽壳黑色素对Pb2+、Cr3+和Cu2+的脱除效率
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