皱瘤海鞘抗人肺癌A549细胞组分的分离纯化及化学成分分析  

Purification and Composition Analysis of Antitumor Compounds from Styela plicata against Lung Cancer A549 Cells

作　　者：毛楷林 林芳 徐腾 余作挺 周海龙 MAO Kailin;LIN Fang;XU Teng;YU Zuoting;ZHOU Hailong(Institute of Tropical Agriculture and Forestry,Hainan University,Haikou 570228,China)

机构地区：[1]海南大学热带农林学院,海南海口570228

出　　处：《食品科学》2018年第21期196-202,共7页Food Science

基　　金：海南省研究生创新科研课题(Hys2017-67);大学生创新创业训练计划项目(201510589039;201610589033)

摘　　要：采用四氮甲唑蓝比色法检测皱瘤海鞘(Styela plicata)被囊及内脏团甲醇提取物对体外培养的人肺癌A549细胞增殖的抑制作用,利用倒置显微镜、流式细胞仪及荧光显微镜观察,分析其对A549细胞的形态和细胞凋亡的影响,并且用气相色谱-质谱联用(gas chromatography-mass spectrometry,GC-MS)技术分析高活性组分6号内脏团柱层析组分(column eluted fraction 6 from visceral methanol extract,VCEF-6)的化学成分。结果表明:在低质量浓度(25～75μg/m L)范围内,被囊及内脏团醇提物对A549细胞生长没有抑制作用,当质量浓度增大到100μg/m L以上时,被囊及内脏团醇提物表现出对A549细胞明显的生长抑制作用。醇提物柱层析组分的体外抗肿瘤检测表明,共有8个组分在100μg/mL时对A549细胞表现出明显的生长抑制作用,其中VCEF-6抑制A549细胞生长活性最强,24 h时半数抑制浓度为168.7μg/m L。通过流式细胞术以及荧光显微镜观察表明,VCEF-6抑制肿瘤细胞生长与诱导细胞凋亡有关。GC-MS分析VCEF-6的化学成分结果表明,VCEF-6含有22种已知物质,其中甾醇类6种、烷烃类6种、脂肪酸酯类6种、芳香族化合物4种。综上所述,在体外实验中VCEF-6能够抑制A549细胞生长并诱导其凋亡。To evaluate the antitumor efficacy of the methanol extracts from the tunic and viscera of Styela plicata,the inhibitory effect on proliferation of lung cancer A549 cells was measured by methyl thiazolyl tetrazolium assay and the effect on cell morphology and apoptosis was examined using an inverted microscope,a fluorescence-activated cell sorter and a fluorescence microscope.Besides,the chemical composition of potent antitumor fractions was analyzed by gas chromatography-mass spectrometry(GC-MS).The results showed that the methanol extracts from the tunic and viscera of Styela plicata could inhibit the growth of human lung cancer A549 cells at a concentration equal to or greater than 100μg/mL but not at lower concentrations(25–75μg/mL).In vitro tests showed that 8 column chromatographic fractions significantly inhibiting the growth of A549 cells were separated from the methanol extracts and among these,column eluted fraction 6 from visceral methanol extract(VCEF-6)separated from the visceral methanol extract exhibited the strongest antitumor activity with a half inhibition concentration of 168.7μg/mL at 24 h.Further observation and analysis using flow cytometry and fluorescence microscope showed that the inhibitory effect of VCEF-6 on cell proliferation was related to induction of cell apoptosis.GC-MS analysis showed that VCEF-6 contained 22 known compounds,including 6 sterols,6 alkanes,6 aliphatic acid esters and 4 aromatic compounds.In conclusion,VCEF-6 has an inhibitory effect on the proliferation of A549 cells and induces cell apoptosis in vitro.

关 键 词：皱瘤海鞘 抗肿瘤活性 细胞凋亡 气相色谱-质谱联用 

分 类 号：R285.5[医药卫生—中药学]