柠檬皮多酚成分分析及其对胰岛素抵抗HepG2细胞糖代谢的影响

王瑞雪; 张筠; 崔艳伟; 付红岩; 房一明; 张彦军; 初众

doi:10.13386/j.issn1002-0306.2022030281

柠檬皮多酚成分分析及其对胰岛素抵抗HepG2细胞糖代谢的影响

doi: 10.13386/j.issn1002-0306.2022030281

王瑞雪^{1, 2,},
张筠^2, ,,
崔艳伟²,
付红岩²,
房一明¹,
张彦军¹,
初众^1, ,

1.
中国热带农业科学院香料饮料研究所，海南万宁 571533
2.
黑龙江东方学院食品工程学院，黑龙江哈尔滨 150066

基金项目: 农业农村部香辛饮料作物遗传资源利用重点实验室、海南省香料饮料作物工程技术研究中心开放课题资助（2019xys003）；黑龙江东方学院科研创新团队建设项目（HDFKYTD202104）。

详细信息

作者简介:
王瑞雪（1998−），女，硕士研究生，研究方向：食品功能性，E-mail：1186615232@qq.com

通讯作者:
张筠（1976−），女，博士，教授，研究方向：食品功能性，E-mail：forever19@sina.com

初众（1981−），男，硕士，研究员，研究方向：食品工程，E-mail：cz809@163.com

中图分类号: TQ283
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- 被引次数: 0
出版历程
- 收稿日期: 2022-03-23
- 网络出版日期: 2022-10-21
- 刊出日期: 2022-11-23

Analysis of Polyphenols from Lemon Peel and Its Effect on Glucose Metabolism in Insulin-resistant HepG2 Cells

WANG Ruixue^{1, 2
,},
ZHANG Yun^{2
, ,},
CUI Yanwei²,
FU Hongyan²,
FANG Yiming¹,
ZHANG Yanjun¹,
CHU Zhong^{1
, ,}

1.
Chinese Academy of Tropical Agricultural Sciences, Spice and Beverage Research Institute, Wanning 571533, China
2.
College of Food Engineering, East University of Heilongjiang College, Harbin 150066, China

摘要

摘要: 目的：探究柠檬皮多酚（Limon Peel Polyphenols，LPP）的组成成分，并研究其对胰岛素抵抗（Insulin Resistance，IR）的HepG2细胞糖代谢的影响。方法：采用高效液相色谱串联四级杆飞行时间质谱法（HPLC-QTOF-MS）分析LPP组成，利用HepG2细胞建立胰岛素抵抗模型，用LPP作用于IR-HepG2细胞，通过测定细胞葡萄糖消耗量初步探究LPP对糖代谢的影响，再通过测定糖原含量和己糖激酶（Hexokinase，HK）、丙酮酸激酶（Pyruvate Kinase，PK）、磷酸烯醇式丙酮酸羧基酶（Phosphoenol Pyruvate Carboxykinase，PEPCK）、葡萄糖六磷酸酶（Glucose-6-Phosphatase，G6Pase）的活性，探究LPP调节细胞糖代谢的作用途径。结果：经过HPLC-QTOF-MS分析出12种物质，主要为黄酮及其苷类成分；在糖代谢研究方面，与模型组相比浓度为0.1~2 mg/mL柠檬皮多酚可显著提高细胞葡萄糖消耗量（P<0.05），且当浓度为0.5 mg/mL时其提高IR-HepG2细胞糖原含量和HK、PK活性，降低PEPCK和G6Pase活性的能力最接近于二甲双胍阳性对照。结论：柠檬皮多酚可以缓解IR-HepG2细胞的胰岛素抵抗状态，并能通过促进糖原合成、提高糖酵解关键酶活性、降低糖异生酶活力的方式调节糖代谢水平，为后续的体内研究提供了数据支持，并为未来开发功能性产品提供了理论依据。
- 柠檬皮多酚 /
- 成分分析 /
- 降糖活性 /
- HepG2细胞 /
- 胰岛素抵抗
Abstract: Objective: To explore the composition of limon peel polyphenols (LPP) and its influence on glucose metabolism in insulin resistance (IR) HepG2 cells. Methods: The composition of LPP was analyzed by HPLC-QTOF-MS. HepG2 cells were used to establish insulin resistance model, IR-HepG2 cells were treated with LPP, and the effect of LPP on glucose metabolism was preliminarily investigated by measuring glucose consumption. By determining glycogen content and hexokinase (HK), pyruvate kinase (PK), phosphoenol pyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) activities to explore the pathway of LPP regulating glucose metabolism in cells. Results: Twelve compounds were identified by HPLC-QTOF-MS, mainly flavonoids and their glycosides. In the study of glucose metabolism, compared with the model group, the concentration of 0.1~2 mg/mL lemon peel polyphenols significantly increased glucose consumption (P<0.05), and the ability to increase the glycogen content and HK and PK activities and decrease the PEPCK and G6Pase activity of IR-HepG2 cells was closest to metformin-positive control when the concentration of lemon peel polyphenols was 0.5 mg/mL. Conclusion: Lemon peel polyphenols can reduce IR-HepG2 cell state of insulin resistance, and can promote glycogen synthesis, improve the glycolytic key enzyme activity, reduce sugar dysplasia regulate sugar metabolism enzyme activity way, it provides data support for the subsequent research in the body, and provides the theoretical basis for the future development of functional products.
- lemon peel polyphenols /
- component analysis /
- hypoglycemic activity /
- HepG2 cells /
- insulin resistance

HTML全文

图 1 柠檬皮多酚总离子流色谱图

Figure 1. TIC chromatography of lemon peel polyphenols

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图 2 柠檬皮多酚高效液相色谱图

Figure 2. HPLC chromatography of lemon peel polyphenols

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图 3 峰9（b）的EIC图（A）、一级质谱图（B）和二级质谱图（C）

Figure 3. EIC spectra (A)、MS spectra (B) and MS/MS spectra (C) of peak 9(b)

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图 4 LPP对IR-HepG2细胞葡萄糖消耗量的影响

Figure 4. Effect of LPP on the glucose consumption of IR-HepG2 cells

注：相同培养时间条件下，#表示与模型组相比差异显著（P<0.05）；△表示与阳性对照组相比差异显著（P<0.05），未标注表示差异不显著（P>0.05）。

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图 5 LPP对IR-HepG2细胞存活率的影响

Figure 5. Effect of LPP on survival rate of IR-HepG2 cells

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图 6 LPP对IR-HepG2细胞中糖原含量的影响

Figure 6. Effect of LPP on the glycogen content in IR-HepG2 cells

注：与空白对照组相比，*表示P<0.05，**表示P<0.01，***表示P<0.001；与模型组相比，###表示P<0.001；与阳性对照组相比，^△△表示P<0.01，^△△△表示P<0.001；未标记表示P>0.05，图7~图10同。

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图 7 LPP对IR-HepG2细胞中HK活性的影响

Figure 7. Effect of LPP on the activity of HK in IR-HepG2 cells

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图 8 LPP对IR-HepG2细胞中PK活性的影响

Figure 8. Effect of LPP on the activity of PK in IR-HepG2 cells

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图 9 LPP对IR-HepG2细胞中PEPCK活性的影响

Figure 9. Effect of LPP on the activity of PEPCK in IR-HepG2 cells

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图 10 LPP对IR-HepG2细胞中G6Pase活性的影响

Figure 10. Effect of LPP on the activity of G6Pase in IR-HepG2 cells

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表 1 LPP成分分析结果

Table 1. The results of LPP component analysis

序号	保留时间（min）	母离子	子离子	分子式	误差	名称	参考文献
1	3.34	169.0495	93 [M+H-甲氧基-CO-羟基]⁺	C₈H₈O₄	0.06	香草酸	[18]
2	4.93	355.1024	193 [M+H-葡萄糖]⁺	C₁₆H₁₈O₉	−0.12	新绿原酸	[19]
3	7.70	625.1760	487 [M+H-138]⁺	C₂₈H₃₂O₁₆	0.56	Chrysoeriol-7-O-（2'-O-mannopyranosyl）allopyranoside	−
4	7.93	449.1080	329 [M+H-120]⁺	C₂₁H₂₀O₁₁	−0.21	荭草素	[20]
5	9.18	433.1131	283 [M+H-阿拉伯糖]⁺	C₂₁H₂₀O₁₀	−0.85	山奈黄苷	[21]
6	9.19	597.1815	289 [M+H-新橙皮糖]⁺	C₂₇H₃₂O₁₅	−0.27	圣草枸橼苷	[22]
7	9.37	579.1708	271 [M+H-新橙皮糖]⁺， 433 [M+H-鼠李糖]⁺	C₂₇H₃₀O₁₄	−0.56	野漆树苷	[17]
8	10.26	463.1233	313 [M+H-阿拉伯糖]⁺	C₂₂H₂₂O₁₁	−0.4	金雀花素	−
9（a）	11.94	609.1813	301 [M+H-新橙皮糖]⁺， 463 [M+H-鼠李糖]⁺	C₂₈H₃₂O₁₅	−0.86	地奥司明	[23]
9（b）	11.98	611.1960	303 [M+H-鼠李糖-葡萄糖]⁺， 465 [M+H-鼠李糖]⁺	C₂₈H₃₄O₁₅	0.39	橙皮苷	[17]
10	12.41	353.0855	147 [M+H-206]⁺	C₁₆H₁₆O₉	0.61	4-甲基-7-乙酰氧基香豆素-β-D-葡萄糖醛酸苷	−
11	24.59	515.2277	161 [M+H-C₁₈H₂₈O₈]⁺， 411 [M+H-C₃H₄O₄]⁺	C₂₈H₃₄O₉	−0.2	诺米林	[24]

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