Optimization of Aqueous Extraction Preparation Technology of Cistanche deserticola by Response Surface Methodology and Its Activity Research
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摘要: 为了研究肉苁蓉水提物制备工艺及其营养价值和活性功能研究，本实验采用水提取法，考察不同提取条件对肉苁蓉浸膏得率的影响，并进一步采用响应面法优化工艺，再测定提取物营养成分及功效成分含量，讨论其抗氧化功能及对秀丽隐杆线虫寿命和生殖力的影响。结果显示，当料液比为1:12 g/mL，煎煮时间为140 min，煎煮次数为4次，肉苁蓉水提物（Aqueous extract of Cistanche, AEC）浸膏得率为65.83%±0.87%。此外，检测AEC的营养成分和活性成分，发现其含有大量的营养成分，例如蛋白质（5.22±0.44 g/100 g）、脂肪（0.150±0.06 g/100 g）、氨基酸（0.640±0.05 g/100 g）等，以及松果菊苷（557±4.24 mg/100 g）、毛蕊花糖苷（169±7.07 mg/100 g）、总糖（22.1±0.64 g/100 g）、总黄酮（4.13±0.08 g/100 g）等多种功效成分。AEC还具有良好的DPPH·清除能力、抑制·OH能力、总抗氧化能力和还原能力，且均呈浓度依赖性。最后，AEC还可以延长秀丽隐杆线虫的寿命，其中8 mg/mL AEC效果最好，平均寿命可达10.95±2.40 d。由此可见，水提取后的AEC具有良好的抗氧化能力，富含多种营养成分和功效成分，这提示肉苁蓉具备开发食品抗氧化剂或功能食品的潜力，为日后肉苁蓉的临床应用研究提供一定的理论依据。Abstract: To study the nutritional value and antioxidant activities of Cistanche deserticola Y. C. Ma extract. The water extraction method was used in this experiment to research the effects of different extraction conditions on the dry extract rate of Cistanche deserticola Y. C. Ma. Then the response surface analysis method was further used to optimize the process. Moreover, the nutritional components and functional components of the extract were determined, and the antioxidant function and effects on the life span and fecundity of C. elegans were discussed. The results showed that when the material liquid ratio was 1:12 g/mL, the decocting time was 140 min, and the decocting times were 4 times, the dry extract yield of the aqueous extract of Cistanche (AEC) was 65.83%±0.87%. In addition, the nutritional composition and functional components of AEC was detected and found to contain a large amount of nutritional components such as protein (5.22±0.44 g/100 g), fat (0.150±0.06 g/100 g), amino acids (0.640±0.05 g/100 g), etc, and echinacoside (557±4.24 mg/100 g), acteoside (169±7.07 mg/100 g), total sugar (22.1±0.64 g/100 g), total flavonoids (4.13±0.08 g/100 g) and others. Furthermore, the AEC also showed good DPPH scavenging activity, hydroxyl radical scavenging activity, total antioxidant activity, and reducing activity, all of which were concentration dependent. Finally, AEC could prolong the life span of Caenorhabditis elegans. Among them, 8 mg/mL AEC had the best effect, and its average life span reached 10.95±2.40 d. Therefore, AEC has good antioxidant capacity, and rich in various nutrients and functional components, which indicates that Cistanche deserticola Y. C. Ma has the potential to develop functional food antioxidants and provides a theoretical basis for the clinical application of Cistanche deserticola Y. C. Ma in the future.
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图 3 不同煎煮时间对AEC浸膏得率的影响
Figure 3. Effect of different decocting time on the yield of AEC
图 4 不同煎煮次数对AEC浸膏得率的影响
Figure 4. Effect of different times of decocting on the yield of AEC
图 5 料液比、煎煮时间和煎煮次数对AEC浸膏得率的等高线图和响应面图
Figure 5. Contour map and response surface map of material to liquid ratio, decocting time and times of decocting on the yield of AEC
图 6 不同浓度AEC总抗氧化能力
Figure 6. Total antioxidant capacity of AEC at different concentrations
图 7 不同浓度AEC清除DPPH自由基的能力
Figure 7. The ability of different concentrations of AEC to scavenge DPPH radicals
图 8 不同浓度AEC清除羟基自由基能力
Figure 8. Hydroxyl radical scavenging ability of AEC at different concentrations
图 10 不同浓度AEC对线虫寿命的影响
Figure 10. Effects of different concentrations of AEC on C. elegans lifespan
图 11 不同浓度AEC对线虫生殖力的影响
Figure 11. Effects of different concentrations of AEC on C. elegans reproductive capacity
注：相同字母代表差异不显著（P>0.05）。
表 1 响应面试验因素水平编码表
Table 1. Coding table of response surface test factor level
序号 因素 −1 0 1 A 料液比（g/mL） 1:10 1:12 1:14 B 煎煮时间（h） 1.5 2.0 2.5 C 煎煮次数（times） 3 4 5 
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表 2 响应面试验设计与结果
Table 2. Response surface test design and results
试验号 A料液比 B煎煮时间 C煎煮次数 浸膏得率（%） 1 −1 0 1 63.10 2 0 0 0 60.09 3 0 −1 1 64.96 4 1 −1 0 58.66 5 0 0 0 61.30 6 0 0 0 58.90 7 0 1 −1 52.02 8 0 −1 −1 39.66 9 1 0 1 65.10 10 −1 −1 0 56.51 11 0 1 1 65.30 12 1 1 0 62.70 13 1 0 −1 41.31 14 −1 0 −1 39.54 15 0 0 0 59.70 16 0 0 0 58.90 17 −1 1 0 61.50
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表 3 二次多项方程的方差分析
Table 3. Analysis of variance of quadratic multinomial equation
来源 平方和 自由度 均值 F值 P值 显著性 模型 1194.57 9 132.73 60.80 <0.0001 ** A 6.34 1 6.34 2.90 0.1322 B 59.02 1 59.02 27.04 0.0013 ** C 923.00 1 923.00 422.8 <0.0001 ** AB 0.23 1 0.23 0.10 0.7572 AC 0.013 1 0.013 0.006058 0.9401 BC 36.12 1 36.12 16.55 0.0048 ** A2 10.50 1 10.50 4.81 0.0644 B2 11.37 1 11.37 5.21 0.0564 C2 148.39 1 148.39 67.97 <0.0001 ** 残差 15.28 7 2.18 失拟项 11.32 3 3.77 3.81 0.1145 ns 总和 1209.86 16 R2：0.9874 C.V.%：2.59 R2Adj：0.9711 R2Pred：0.8452 注：*代表存在显著性差异（P<0.05）；**代表存在极显著性差异（P<0.01）；ns（not significant）代表差异不显著。
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表 4 AEC主要营养成分检测结果
Table 4. Detection of main nutrients in AEC
指标 含量（g/100 g） 蛋白质 5.22±0.44 氨基酸 0.64±0.05 脂肪 0.15±0.06 灰分 2.70±0.00 钠（mg/100 g） 225±3.54 碳水化合物 33.60±1.63 能量（kJ/100 g） 665.50±18.17
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表 5 AEC主要活性成分检测
Table 5. Detection of main active components in AEC
指标 含量（mg/100 g） 松果菊苷 557±4.24 毛蕊花糖苷 169±7.07 粗多糖（g/100 g） 1.86±0.03 京尼平苷酸 13.40±0.64 8-表番木鳖酸 131±3.54 肉苁蓉苷A 126±0.00 管花苷A 20.40±0.21 异类叶升麻苷 194±0.71 2-乙酰基洋丁香氛苷 234±11.31 管花苷B 487±9.90 总糖（g/100 g） 22.10±0.64 总黄酮（g/100 g） 4.13±0.08 尿囊素 293±4.95
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表 6 不同浓度AEC对线虫平均寿命的影响
Table 6. Effects of different concentrations of AEC on C. elegans average lifespan
组别 平均寿命（d） 对照组 9.46±2.69 4 mg/mL 9.88±2.97 8 mg/mL 10.95±2.40** 16 mg/mL 10.05±2.45 注：**表示与对照组存在极显著性差异（P<0.01）。
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