荞麦中光毒物质的转化及荞麦光敏素对人结肠癌细胞HCT-116的抑制活性

Transformation of Phototoxicants from Buckwheat and Inhibitory Activity of Fagopyrins on the Human Colon Cancer Cell Line HCT-116

荞麦光敏素(fagopyrins)具有光毒性和酪氨酸激酶抑制活性,可作为光动力治疗中的光敏剂,用于癌症的治疗。本研究采用电喷雾质谱法和荧光光度法,探究了荞麦中光毒物存在的型式,并分析了光敏素前体(protofagopyrin)转化为荞麦光敏素的条件。通过MTT细胞实验,研究荞麦光敏素提取物对人结直肠癌细胞和人结肠上皮细胞生长增殖的影响。结果表明,荞麦中光毒物以光敏素前体为主,并且有微量的荞麦光敏素,前者质谱的离子强度是后者的400倍。光敏素前体的转化发生在光照提取过程中,而且提取液在光照放置过程中,光敏素前体继续转化,尤其是前0. 5 h对于转化的影响显著超过了提取过程。荞麦光敏素提取物与细胞作用,避光培养时,对HCT-116癌细胞有轻度的抑制作用;而对FHC正常细胞不仅未见抑制还有促进作用。光照结合避光培养时,荞麦光敏素对癌细胞的抑制随着提取物浓度的增加而增强,当提取物浓度达到200μg/m L时,抑制率达到50%;而对正常细胞提取物,浓度低于200μg/m L时未见抑制,500～1 000μg/m L时抑制率达到50%。光敏素提取物与金丝桃素抑制细胞的能力相比,光照结合避光培养时,金丝桃素对正常细胞的抑制作用强于对癌细胞,而荞麦光敏素的提取物正好相反。提示荞麦光敏素比金丝桃素对癌细胞的抑制更具特异性。上述结果为从荞麦叶中开发出有利于肿瘤治疗的药物提供了实验依据。

Fagopyrins are naturally occurring substances in buckwheat plants. They possess strong phototoxicity and tyrosine kinase inhibitory activities, and thus can be utilized as photosensitizers in photodynamic therapy against various types of cancers. In the current study, we investigated the types of phototoxicants from the buckwheat and analyzed the conditions of transformation from protofagopyrin into fagopyrins by electrospray ionization mass spectrometry and fluorescence spectroscopy. We further evaluated the effects of fagopyrins on growth and proliferation of both the human colon cancer cells (HCT-116) and human fetal colon epithelial cells (FHC) by using MTT assays. We found that the phototoxicants contained in the buckwheat were primarily present in the protofagopyrin form with ionic intensity of about 400 times more in contrast to the fagopyrins. The protofagopyrin into fagopyrins transformation was observed during the extraction process in the presence of light, and the conversion greatly exceeded the extraction process during the first 0.5 h of the photostage. Secondly, in contrast to the normal FHC cells, minor inhibitory effects were noticed on HCT-116 cancer cells when exposed to fagopyrins in the dark, which was increased (about 50%) by introducing the light of 1.5 h and by increasing the concentration of the extract up to 200 μg/mL. However, there was no inhibition on FHC when the concentration of the extract was lower than 200 μg/mL, and the inhibition rate was reached about 50% at 500-1 000 μg/mL. In comparison, hypericin has exerted strong inhibitory effects on FHC rather than on HCT-116, while the fagopyrins extracts do the opposite. In summary, this study concluded that the fagopyrin extracts are more specific than hypericin for the inhibition of proliferation of colon cancer cells, and the buckwheat could be a potential source of a wide variety of natural products for the development of anti-cancer drugs.