灰树花不同浓度醇提物的活性成分分析及抗氧化、抗EV71病毒的作用研究

熊雯宇; 何君强; 汤丹琳; 彭培植; 江小琴; 刘斌

灰树花不同浓度醇提物的活性成分分析及抗氧化、抗EV71病毒的作用研究

熊雯宇^1,,
何君强¹,
汤丹琳¹,
彭培植²,
江小琴¹,
刘斌^{1, 2,}

1.
福建农林大学食品科学学院，福建　福州　350002
2.
国家菌草工程技术研究中心，福建　福州　350002

基金项目: 中国博士后科学基金（2020M671918）；福建省科学技术厅区域发展项目（2020N3002）；福建省科学技术厅对外合作项目（2021I0008）；福建省科技重大专项（2021NZ0101）

详细信息

作者简介:
熊雯宇（ 1996−），女，硕士研究生．研究方向：食药用菌活性成分及功能营养（E-mail：1184503923@qq.com）

通讯作者:
刘斌( 1966−) ，男，博士，教授，研究方向: 食品生物技术、分子营养学（E-mail：liubin618@ hotmail.com）

中图分类号: TS255.3
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出版历程
- 收稿日期: 2022-12-02
- 修回日期: 2022-12-22
- 网络出版日期: 2023-04-14

Study on the Active Ingredient and Their Antioxidative and Anti-EV71 Virus Effects in Different Concentrations of Ethanol Extract from Grifola frondosa

1.
College of Food Science, Fujian Agriculture and Forestry University, Fuzhou, Fujian　　350002, China
2.
National Engineering Research Center of JUNCAO Technology, Fuzhou, Fujian　　350002, China

摘要

摘要: 目的探究灰树花适宜的乙醇提取浓度，分析不同乙醇浓度提取对灰树花活性成分及抗氧化、抗EV71病毒活性的影响，促进灰树花的开发利用。方法分别采用55%、70%和95%的乙醇溶液作为提取溶剂对灰树花子实体进行超声波辅助提取，测定不同浓度乙醇提取物主要活性成分含量并对其抗氧化、抗EV71病毒能力进行相关性分析。结果不同乙醇浓度提取物活性成分含量存在差异，灰树花95%乙醇提取物（GFE95）中多酚和三萜含量最高分别为0.042%和1.82%，灰树花55%乙醇提取物（GFE55）中多糖和蛋白质含量最高，分别为33.08%和17.45%。GFE55、灰树花70%乙醇提取物（GFE70）和GFE95具有不同程度的抗氧化和EV71病毒抑制作用。其中，GFE95的抗氧化能力最好，对2,2-联苯基-1-苦基肼基（DPPH）、2,2′-联氨双二胺盐（ABTS⁺）、超氧阴离子（O₂⁻）和过氧化氢（H₂O₂）自由基的清除能力的IC₅₀值分别为0.7427、0.7414、0.902、0.4146 mg·mL⁻¹。GFE95的抗病毒作用较强，当样品质量浓度为200 μg·mL⁻¹时，GFE95对EV71病毒的抑制率最高，达88.18%，其对EV71病毒抑制率的IC₅₀值为194.80 μg·mL⁻¹，TI值为9.65。皮尔逊相关性分析结果表明DPPH、ABTS⁺、O₂⁻自由基清除率与三萜类成分含量呈极显著正相关，相关系数分别为0.992、0.971和0.613（P＜0.01）。O₂⁻、H₂O₂自由基清除率与多酚类成分含量呈极显著正相关，相关系数分别为0.921和0.997（P＜0.01）。EV71病毒抑制作用与多酚、三萜类成分含量呈极显著正相关（P＜0.01），相关系数分别为0.536和0.954。灰树花中三萜和多酚的含量与抗氧化、抗病毒活性具有较强的相关性。结论以体积分数为95%的乙醇溶液为提取剂，可有效提取灰树花中具有抗氧化、抗病毒活性的多酚、三萜、多糖等活性物质，提高灰树花的生物活性，是进一步促进灰树花的高值化应用开发的关键。研究结果为灰树花的提取溶剂选择及活性作用的研究奠定基础。
- 灰树花 /
- 乙醇提取 /
- 活性成分 /
- 抗氧化作用 /
- 抗病毒作用
Abstract: Objective In order to explore the appropriate ethanol extraction concentration of Grifola frondosa, analyze the effects of different ethanol concentrations on the active components and antioxidant and anti-EV71 virus activities of G. frondosa, and promote the development and utilization of G. frondosa. Method The 55%, 70% and 95% ethanol solutions were used as solvent for ultrasonic-assisted extraction of G.frondosa fruiting bodies. The contents of main active components in different concentrations of ethanol extracts and their antioxidant and anti-EV71 virus abilities were determined for correlation analysis. Result There were differences in the content of active ingredients in different ethanol concentration extracts. The highest contents of polyphenols and triterpenes in 95% ethanol extract of G. frondosa (GFE95) were 0.042% and 1.82%, respectively. The highest contents of polysaccharides and proteins in 55% ethanol extract of G. frondosa (GFE55) were 33.08% and 17.45%, respectively. GFE55, 70% ethanol extract of G. frondosa (GFE70) and GFE95 have different degrees of antioxidant and EV71 virus inhibition. Among them, GFE95 had the best antioxidant capacity, and its IC₅₀ values for scavenging DPPH, ABTS⁺, O₂^- and H₂O₂ free radicals were 0.7427 mg/mL, 0.7414 mg/mL, 0.9026 mg/mL and 0.4146 mg/mL, respectively. GFE95 had a strong antiviral effect. When the sample concentration was 200 μg/mL, the inhibition rate of GFE95 on EV71 virus reached 88.18%. The IC₅₀ value of GFE95 on EV71 virus inhibition rate was 194.80 μg/mL, and the TI value was 9.65. Pearson’s correlation analysis showed that DPPH, ABTS⁺ and O₂^- free radical scavenging rates were significantly positively correlated with the content of triterpenoids, with correlation coefficients of 0.992, 0.971 and 0.613 (P＜0.01), respectively. O₂^- and H₂O₂ free radical scavenging rates were significantly positively correlated with the content of polyphenols, with correlation coefficients of 0.921 and 0.997 (P＜0.01), respectively. The inhibitory effect of EV71 virus was significantly positively correlated with the content of polyphenols and triterpenoids (P＜0.01), and the correlation coefficients were 0.536 and 0.954, respectively. The contents of triterpenoids and polyphenols in G. frondosa had strong correlation with antioxidant and antiviral activities. Conclusion Using 95% ethanol solution as the extractant, the active substances such as polyphenols, triterpenes and polysaccharides with antioxidant and antiviral activities in G. frondosa can be effectively extracted, and the biological activity of G. frondosa can be improved, which is the key to further promote the high-value application and development of G. frondosa. The results lay a foundation for the study of extraction solvent selection and activity of G. frondosa.
- Grifola frondosa /
- ethanol extract /
- active ingredient /
- antioxidant activity /
- antiviral activity

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图 1 灰树花不同浓度乙醇提取物中的活性物质含量

不同字母表示处理间差异显著（P＜0.05）。

Figure 1. Contents of active substances in different concentrations of ethanol extract from G. frondosa

Different letters indicate significant differences between treatments (P＜0.05).

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图 2 灰树花不同质量浓度乙醇提取物的DPPH自由基清除率

Figure 2. DPPH free radical scavenging rate in different concentrations of ethanol extract from G. frondosa

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图 3 灰树花不同浓度乙醇提取物的ABTS⁺自由基清除率

Figure 3. ABTS⁺ free radical scavenging rate in different concentrations of ethanol extract from G. frondosa

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图 4 灰树花不同浓度乙醇提取物的O₂^-自由基清除率

Figure 4. O₂^- free radical scavenging rate in different concentrations of ethanol extract from G. frondosa

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图 5 灰树花不同浓度乙醇提取物的H₂O₂自由基清除率

Figure 5. H₂O₂ free radical scavenging rate in different concentrations of ethanol extract from G. frondosa

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图 6 灰树花不同浓度乙醇提取物对Vero细胞的物质毒性

与浓度0 μg·mL⁻¹相比， **代表P＜0.01；

Figure 6. Substance toxicity to Vero cells in different concentrations of ethanol extract from G. frondosa

Compared with the 0 μg·mL⁻¹, ** representd P＜0.01 ;

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图 7 灰树花不同浓度乙醇提取物对EV71病毒的抑制率

Figure 7. Inhibition rate of EV71 virus in different concentrations of ethanol extract from G. frondosa

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表 1 灰树花不同浓度乙醇提取物抗氧化作用的IC₅₀值

Table 1. IC₅₀ Value of antioxidation in different concentrations of ethanol extract from G. frondosa

样品名称 description of sample	DPPH自由基清除能力IC₅₀值 DPPH·/( mg·mL⁻¹)	ABTS阳离子自由基清除能力IC₅₀值 ABTS⁺·/( mg·mL⁻¹)	超氧阴离子自由基清除能力IC₅₀值 O₂⁻·/( mg·mL⁻¹)	过氧化氢自由基清除能力IC₅₀值 H₂O₂·/( mg·mL⁻¹)
GFE55	1.3070	0.9327	1.1960	1.0140
GFE70	0.9958	0.8085	1.4280	2.5830
GFE95	0.7427	0.7414	0.9026	0.4146

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表 2 灰树花不同浓度乙醇提取物对EV71病毒的TC₅₀ 、TC₅₀ 和TI

Table 2. TC₅₀, TC₅₀ and TI of different concentrations of ethanol extract from G. frondosa on EV71 virus

样品名称 Description of sample	样品半数中毒浓度 TC₅₀/(μg·mL⁻¹)	抗EV71病毒半数有效浓度 IC₅₀/(μg·mL⁻¹)	治疗指数 TI
GFE55	1868.00	440.60	4.24
GFE70	1869.00	401.50	4.66
GFE95	1879.00	194.80	9.65

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表 3 灰树花不同浓度乙醇提取物活性物质的含量与抗氧化、抗EV71病毒活性IC₅₀的相关系数

Table 3. The correlation coefficients between the IC₅₀ of antioxidant and anti-EV71 activities and the contents of active substance in different concentrations of ethanol extract from G. frondosa

指标 Index	多酚 Polyphenols	三萜 Triterpenoids	多糖 Polysaccharide	蛋白质 Protein
DPPH自由基清除率IC₅₀ IC₅₀ of DPPH radical scavenging rate	0.130	0.992**	−0.252	−0.997**
ABTS⁺自由基清除率IC₅₀ IC₅₀ of ABTS⁺ radical scavenging rate	0.019	0.971**	−0.358	−0.982**
O₂⁻自由基清除率IC₅₀ IC₅₀ of O₂⁻ radical scavenging rate	0.921**	0.613**	0.706**	−0.573**
H₂O₂自由基清除率IC₅₀ IC₅₀ of H₂O₂ radical scavenging rate	0.997**	0.334	0.893**	−0.286
EV71病毒抑制率IC₅₀ IC₅₀ of EV71 virus inhibition rate	0.536**	0.954**	0.179	−0.937**
注：代表显著相关（P＜0.05），代表极显著相关（P＜0.01）； Note: representd significant correlation (P＜0.05), ** representd extremely significant correlation (P＜0.01) ;

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