目的 分離純化制備羊棲菜褐藻糖膠（CSFP-1），并對其抗腫瘤活性及可能作用機制進行初步研究。方法 通過冷水浸提法制備羊棲菜褐藻糖膠（CSFP），經(jīng)超濾系統(tǒng)分離獲得CSFP-1，并采用苯酚-硫酸比色法、PMP衍生化、高效液相色譜法、紅外光譜分析等多種方法對其理化性質(zhì)進行分析；采用MTT法評價CSFP-1（0、10、100、300 μg/mL）對宮頸癌細胞（HeLa）、肺癌細胞（A549）和肝癌細胞（Hep3B）生存能力的影響；應(yīng)用流式細胞術(shù)評價CSFP-1對HeLa細胞細胞周期的影響；應(yīng)用Western blotting技術(shù)評價CSFP-1對HeLa細胞凋亡和自噬相關(guān)蛋白表達的影響。結(jié)果 CSFP-1的糖含量為84.18%，糖醛酸含量為14.28%，硫酸根含量為10.02%，平均相對分子質(zhì)量為5.6×10⁴，主要由半乳糖、巖藻糖、甘露糖、葡萄糖醛酸和木糖組成。CSFP-1具有選擇性細胞毒性，對HeLa細胞生長發(fā)揮顯著抑制作用（P<0.05、0.001），對A549和Hep3B細胞存活率沒有明顯影響。與對照組比較，HeLa細胞經(jīng)CSFP-1處理24 h后，G1期細胞比例明顯增加（P<0.001）；經(jīng)CSFP-1處理48 h后，S期細胞比例顯著增加（P<0.05、0.01、0.001），G2期細胞比例顯著減少（P<0.01、0.001），均呈現(xiàn)劑量相關(guān)性。與對照組比較，CSFP-1 100、300 μg/mL劑量組處理24 h，300 μg/mL劑量組處理48 h Bcl-2蛋白表達顯著減少（P<0.05）；100、300 μg/mL劑量組處理24、48 h Beclin-1蛋白表達顯著增加（P<0.05、0.01），且存在時間和劑量相關(guān)性。結(jié)論 CSFP-1具有選擇性抑制HeLa細胞增殖作用，作用機制可能與阻滯細胞周期、誘導(dǎo)細胞凋亡相關(guān)。

Objective To extract, purify, and prepare the fucoidin of Sargassum fusiforme (CSFP-1), and preliminary investigate its anti-tumor activity and underlying mechanism. Methods CSFP was extracted by cold water and CSFP-1was obtained by ultrafiltration system. Phenol-sulfuric acid colorimetry, PMP derivatization, high performance liquid chromatography, infrared spectroscopy, and other methods were used to analyze the physical and chemical properties. The effects of CSFP-1 (0, 10, 100, and 300 μg/mL) on the viability of cervical cancer cells (HeLa), lung cancer cells (A549), and hepatoma cells (Hep3B) were evaluated by MTT assay, the effects of CSFP-1 on the cell cycle of HeLa were evaluated by flow cytometry, and the effects of CSFP-1 on apoptosis and autophagy related protein expression in HeLa cells were evaluated by Western blotting. Results CSFP-1 was mainly composed of galactose, fucose, mannose, glucuronic acid, and xylose with the sugar content of 84.18%, glucuronic acid content of 14.28%, sulfate content of 10.02% and average molecular weight of 56 kDa. CSFP-1 had selective cytotoxicity and significantly inhibited the growth of HeLa cells (P < 0.05, 0.001), but had no significant effect on the survival rate of A549 and Hep3B cells. Compared with the control group, HeLa cells treated with CSFP-1 for 24 h increased the proportion of G1 phase cells significantly (P < 0.001); After treated with CSFP-1 for 48 h, the proportion of S phase cells increased significantly (P < 0.05, 0.01, 0.001), and the proportion of G2 phase cells decreased significantly (P < 0.01, 0.001). Compared with control group, the expression of Bcl-2 protein in 100 and 300 μg/mL CSFP-1 groups decreased significantly at 24 h, and that in 300 μg/mL group decreased significantly at 48 h (P < 0.05); The expression of Beclin-1 protein in 100 and 300 μg/mL group increased significantly at 24 and 48 h (P < 0.05 and 0.01), and there was a time-dose correlation. Conclusion CSFP-1 has selective inhibited effect on the proliferation of HeLa cells and its anti-tumor activity mechanism may be related with blocking cell cycle and inducing apoptosis.

國家自然科學基金資助項目（314070430）；浙江省教育廳科研項目（Y201737374）；浙江省基礎(chǔ)公益研究計劃（LGN18C020004）；浙江省科技計劃項目（2017C32103）；浙江省自然科學基金（LY18C020006）