摘要
利用DEAE-Sepharose Fast Flow和Sephadex G-200对树莓粗多糖进行分离纯化得到组分RPP-5,探究RPP-5的结构特征和免疫调节活性。采用高效凝胶渗透色谱、傅里叶红外光谱和气相色谱-质谱联用技术对RPP-5的结构进行表征。利用RAW264.7细胞研究RPP-5的免疫活性,通过CCK-8法、微板法、酶联免疫吸附测定和实时荧光定量PCR分别测定RPP-5对RAW264.7细胞的细胞活力、NO含量、炎症因子白细胞介素-6(interleukin-6,IL-6)、白细胞介素-1β(interleukin-1β,IL-1β)和肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)的含量以及炎症因子mRNA表达水平的影响。结果表明,RPP-5主要由物质的量比为44.5∶21.7∶5.9∶8.8∶5∶4.1的阿拉伯糖、葡萄糖、鼠李糖、半乳糖、甘露糖和木糖组成,其重均分子质量、数均分子质量和峰位分子质量分别为7598 Da、6183 Da和7492 Da,共存在12种单糖连接方式,糖链中→4)-Gal p-(1→、→4)-Xyl p-(1→、Ara f-(1→和→4,6)-Glc p-(1→的含量较高,分别为26.27%、16.50%、11.38%和10.41%。RPP-5可显著升高RAW264.7细胞的细胞活力,提高细胞内NO、IL-6、IL-1β和TNF-α的含量,促进IL-6、IL-1β和TNF-α的mRNA表达。综上,RPP-5是一种多支链的酸性杂多糖,具有较强的免疫增强活性,可作为功能性食品中的免疫调节剂,为树莓多糖的结构表征和免疫活性研究提供理论支持。
In the present study,a raspberry polysaccharide named RPP-5 was isolated and purified using DEAE-Sepharose Fast Flow and Sephadex G-200.Its structural characteristics and immunomodulatory activity were also explored.The structural characteristics of RPP-5 were studied based on the results of high-performance gel permeation chromatography,Fourier transform infrared spectroscopy,and GC-MS.RAW264.7 cells were utilized to evaluate the immunomodulatory activity of RPP-5.The cell viability,contents of NO and inflammatory factors,and mRNA expression levels of inflammatory factors in RAW264.7 cells were measured by CCK-8 assay,microplate method,enzyme-linked immunosorbent assay assays,and real-time quantitative polymerase chain reaction,respectively.Results showed that RPP-5 was composed of arabinose,glucose,rhamnose,galactose,mannose,and xylose,with a molar ratio of 44.5∶21.7∶5.9∶8.8∶5∶4.1.The average molecular weight,peak molecular weight,and number average molecular weight were 7598 Da,7492 Da,and 6183 Da,respectively.RPP-5 contained 12 types of glycosyl linkages types,among which→4)-Gal p-(1→,→4)-Xyl p-(1→,Ara f-(1→,→4,6)-Glc p-(1→were the main connection types of sugar residues,accounted for 26.27%,16.50%,11.38%,and 10.41%,respectively.RPP-5 significantly increased the viability of RAW264.7 cells,increased the content of NO,IL-6,IL-1β,and TNF-α,and promoted the expression of IL-6,IL-1β,and TNF-αmRNA in RAW264.7 cells.In conclusion,RPP-5 was a homogeneous acidic heteropolysaccharide with a complex multi-branched structure and showed significant immune-enhancing activity.It could be used as an immunomodulator in functional foods,providing theoretical support for the structural characterization and immunological activity studies of raspberry polysaccharides.
作者
尹星星
杨永晶
陆杰
王学红
晁沐
YIN Xingxing;YANG Yongjing;LU Jie;WANG Xuehong;CHAO Mu(School of Ecological and Environmental Engineering,Qinghai University,Xining 810016,China;Qinghai Raspberry Agricultural Industrialization Co.Ltd.,Xining 812100,China)
出处
《食品与发酵工业》
CAS
CSCD
北大核心
2024年第9期139-147,共9页
Food and Fermentation Industries
基金
青海省科技厅基础研究项目(2019-ZJ-953Q)
国家自然科学基金项目(81860619)。
关键词
树莓
多糖组分
结构特征
免疫活性
RAW264.7细胞
raspberry
polysaccharide
structural characteristics
immunomodulatory activity
RAW264.7 cell
