食品科学 ›› 2020, Vol. 41 ›› Issue (5): 39-48.doi: 10.7506/spkx1002-6630-20190115-165

• 基础研究 • 上一篇    下一篇

‘巨峰’葡萄皮花色苷的分离纯化、结构鉴定及抗肿瘤活性

薛宏坤,谭佳琪,刘钗,刘成海   

  1. (1.东北农业大学工程学院,黑龙江 哈尔滨 150030;2.东北农业大学农学院,黑龙江 哈尔滨 150030)
  • 出版日期:2020-03-15 发布日期:2020-03-23
  • 基金资助:
    东北农业大学“青年才俊”项目(518076)

Isolation, Purification, Structure Identification and Anti-tumor Activity of Anthocyanin from Kyoho Grape Skins

XUE Hongkun, TAN Jiaqi, LIU Chai, LIU Chenghai   

  1. (1. School of Engineering, Northeast Agricultural University, Harbin 150030, China; 2. School of Agriculture, Northeast Agricultural University, Harbin 150030, China)
  • Online:2020-03-15 Published:2020-03-23

摘要: 为研究‘巨峰’葡萄皮花色苷分离纯化方法、结构鉴定及抗肿瘤活性,将葡萄皮色素的粗提物依次经过AB-8大孔树脂、聚酰胺树脂、Sephadex LH-20葡聚糖凝胶进行分离纯化;利用超高效液相色谱三重四极杆飞行时间质谱联用技术对分离所得的花色苷进行结构鉴定;通过噻唑蓝法、Transwell侵袭实验及Annexin V-FITC/PI双标记染色法分别研究纯化后所得的花色苷组分对HepG2肝癌细胞和A549肺癌细胞增殖、侵袭和凋亡能力的影响。结果表明:紫外-可见光谱扫描发现经AB-8大孔树脂纯化后所得的样品溶液(样品1)在280、520、330 nm波长处均有吸收峰,说明样品1中主要成分为花色苷类,同时可能含有酰化基团;样品1再经聚酰胺树脂纯化后发现聚酰胺树脂对花色苷产生吸附作用,而经Sephadex LH-20凝胶纯化后花色苷得到较好的分离,最终得到3 种花色苷组分(色素I、色素III和色素IV),分别为矢车菊-3-芸香糖苷、锦葵素-3,5-双葡萄糖苷-香豆酰和锦葵-3-半乳糖苷,它们所对应的得率和纯度分别为(0.108±0.011)%、(0.053±0.007)%、(0.038±0.005)%和98.52%、96.84%、91.36%;通过3 种花色苷组分对抗肿瘤活性分析发现,3 种花色苷组分均能显著抑制HepG2肝癌细胞和A549肺癌细胞的增殖和侵袭能力,并且能够显著增加癌细胞的凋亡,其抗肿瘤效果顺序依次为色素III>色素I>色素IV。

关键词: 葡萄皮花色苷, 分离, 纯化, 结构, 抗肿瘤作用

Abstract: The purpose of this study was to investigate the isolation, purification, structure and antitumor activity of anthocyanins from kyoho grape skins. Pigment-rich crude extract from kyoho grape skins was separated and purified sequentially by AB-8 macroporous resin, polyamide resin and Sephadex LH-20 column chromatography. The structure of the purified anthocyanins was identified by using ultra-high performance liquid chromatography combined with triple quadrupole time-of-flight mass spectrometry (UPLC-QTOF MSE), and their effects on the proliferation, invasion and apoptosis of HepG2 hepatocellular cells and A549 lung cancer cells were investigated by MTT assay, Transwell invasion assay and Annexin V-FITC/PI double labeling staining, respectively. The ultraviolet (UV)-visible (Vis) spectrum showed that the AB-8 purified sample had absorption peaks at 280, 520 and 330 nm, indicating that the major components of the sample were anthocyanins which might contain acylation groups. In the sample, some anthocyanins were adsorbed by polyamide resin, while Sephadex LH-20 column chromatography could allow good separation and purification of anthocyanins from kyoho grape skins. Three anthocyanin components (I, III and IV) were obtained in high yields ((0.108 ± 0.011)%, (0.053 ± 0.007)% and (0.038 ± 0.005)%) and purities (98.52%, 96.84% and 91.36%, respectively) and they were identified as cyanidin-3-rutinoside, malvidin-3,5-diglucoside-coumarin and malvidin-3-galactoside. The anthocyanin components could significantly inhibit the proliferation and invasion of HepG2 hepatoma cells and A549 lung cancer cells, and significantly increase the apoptosis of cancer cells. Their anti-tumor effects were ranked as follows: III > I > IV.

Key words: grape skin anthocyanins, isolation, purification, structure, anti-tumor effect

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