首页|低相对分子质量海藻多糖的分离纯化、结构鉴定及其生物活性研究

低相对分子质量海藻多糖的分离纯化、结构鉴定及其生物活性研究

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目的 从海藻中制备并分离纯化一种低相对分子质量海藻多糖(LMSP),初步分析其结构特征并研究其抗氧化、降血糖和抗肿瘤作用.方法 水提醇沉法制备海藻粗多糖,H2O2/维生素C(VC)法降解海藻多糖,经DEAE-52阴离子交换色谱柱和葡聚糖凝胶G-100色谱柱分离纯化;采用苯酚-硫酸法测定总糖含量,间羟联苯法测定糖醛酸含量,BaCl2-明胶比浊法测定硫酸基含量,考马斯亮蓝法测定蛋白质含量,高效凝胶渗透色谱测定相对分子质量,进行红外光谱法分析,高效液相色谱测定单糖组成,气相色谱-质谱联用仪检测糖苷键类型,扫描电镜分析形貌特征;1,1-二苯基-2-三硝基苯肼(DPPH)自由基清除率实验、羟自由基清除率实验、2,2-联氮-二(3-乙基-苯并噻唑-6-磺酸)二铵盐(ABTS)自由基清除率实验检测LMSP抗氧化活性;α-淀粉酶抑制实验、α-葡萄糖苷酶抑制实验检测LMSP降血糖活性;检测LMSP对A549、HeLa、Hep-G2细胞的抗肿瘤活性.结果 LMSP的重均相对分子质量为4 635,总糖质量分数为(83.85±2.12)%,糖醛酸的质量分数为(29.73±3.57)%,硫酸根质量分数为(24.44±1.97)%,蛋白质的质量分数为(1.35±0.63)%,主要由半乳糖、木糖和岩藻糖组成,红外光谱及气相色谱-质谱联用对其结构分析推测,该多糖可能是一种含吡喃糖苷的硫酸多糖,主要含有2-Rhap、T-Galp、3-Galp和4-Glcp等片段,LMSP在扫描电镜下呈现出带孔的网状结构,堆叠紧密.LMSP对DPPH自由基清除率、羟基自由基清除率和ABTS自由基清除率的半数抑制浓度(IC50)为1.35、0.61、1.31 mg·mL-1;对α-淀粉酶和α-葡萄糖苷酶的IC50分别为0.82、1.27 mg·mL-1;对 A549、HeLa和 Hep-G2细胞的IC50为2.31、3.41、3.10 mg·mL-1,且各指标的作用均强于海藻粗多糖.结论 H2O2/V C法降解制备得到均一的LMSP,具有良好的抗氧化、降血糖和抗肿瘤作用.
Isolation,purification,structure identification and biological activity of low relative molecular weight Sargassum polysaccharides
Objective To isolate and purify a low molecular weight Sargassum polysaccharides(LMSP)from Sargassum of traditional Chinese medicine,and preliminarily analyze its structural characteristics and study its antioxidant,hypoglycemic and anti-tumor effects.Methods The seaweed polysaccharides were extracted by water extraction and ethanol precipitation,and then degraded by H2O2NC method.They were purified by DEAE-52 anion exchange column and Sephadex G-100 gel filtration column.The total sugar content was determined by phenol-sulfuric acid method,the sugarfarn acid content was determined by p-hydroxybenzaldehyde method,the sulfate content was determined by BaCl2-gelatin turbidimetry method,and the protein content was determined by Coomassie brilliant blue method.The relative molecular mass was determined by high-performance gel permeation chromatography,the infrared spectrum was analyzed,and the composition of monosaccharides was determined by high-performance liquid chromatography.The type of glycosidic bond was detected by gas chromatography-mass spectrometry.The morphological features were analyzed by scanning electron microscopy.The antioxidant activity of LMSP was determined by 1,1-diphenyl-2-picrylhydrazyl(DPPH)free radical scavenging rate experiment,hydroxyl radical scavenging rate experiment,and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)(ABTS)free radical scavenging rate experiment.The hypoglycemic activity of LMSP was determined by α-amylase inhibition experiment and α-glucosidase inhibition experiment.The antitumor activity of LMSP against A549,Hela,and Hep-G2 cells was determined.Results The relative molecular mass of LMSP is 4 635,with a total sugar content of(83.85±2.12)%and a sugar aldonic acid content of(29.73±3.57)%.The sulfate content is(24.44±1.97)%,the protein content is(1.35±0.63)%,and it is mainly composed of galactose,xylose,and fucose.Infrared spectroscopy and gas chromatography-mass spectrometry were used to analyze its structure,and it was inferred that LMSP may be a pyransulfonic polysaccharide containing pyransulfonic glycosides,mainly containing 2-Rhap,T-Galp,3-Galp,and 4-Glcp fragments.Under scanning electron microscope,LMSP presents a porous network structure,tightly stacked.The half maximal inhibitory concentration(IC50)for DPPH radical scavenging,hydroxyl radical scavenging,and ABTS radical scavenging were 1.35,0.61,and 1.31 mg·mL-1,respectively.The IC50 values for α-amylase and α-glucosidase inhibition were 0.82 and 1.27 mg·mL-1,respectively.The IC50 values for A549,HeLa,and Hep-G2 cell inhibition were 2.31,3.41,and 3.10 mg·mL-1,respectively,and the effects of all indicators were stronger than those of seaweed crude polysaccharides.Conclusion The LMSP prepared by the H2O2/V C method is homogeneous and has good antioxidant,glucose-lowering,and anti-tumor effects.

Sargassum polysaccharideslow relative molecular weightH2O2/V C degradationisolation and purificationstructural characterizationbiological activityantioxidanthypoglycemicanti-tumor

朱阳东、丁金铭、于斋卓、尹秋燕、闫滨

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山东中医药大学,山东 济南 250355

海藻多糖 低相对分子质量 H2O2/V C法降解 分离纯化 结构表征 生物活性 抗氧化 降血糖 抗肿瘤

2024

药物评价研究
天津药物研究院 中国药学会

药物评价研究

CSTPCD北大核心
影响因子:1.199
ISSN:1674-6376
年,卷(期):2024.47(12)