灵芝多糖肽GL-PPSQ_2结构研究及其应用

Structure and application of Ganoderma polysaccharide peptide GL-PPSQ_2

目的从灵芝Ganoderma水提物中分离纯化活性多糖肽,对其进行结构研究,并作为对照品用于灵芝产品中多糖肽含量测定。方法采用热水提取、膜超滤技术和凝胶过滤色谱等方法进行分离纯化,根据理化测定及波谱数据分析鉴定化合物结构。检测多糖肽含量采用高效液相色谱仪附紫外检测器,以水作为流动相,体积流量为1mL/min。结果灵芝多糖肽相对分子质量为5.0×10~4,质量分数97%以上,得率为0.49%,多糖含量达87.17%,单糖组成以葡萄糖为主和少量甘露糖组成的多糖,其物质的量比为31.85∶1.00,含有16种氨基酸,氨基酸总量为5.04%。依据甲基化、一维和二维核磁谱图,GL-PPSQ2结构重复单元为主链由→3)-β-D-Glcp-(1→构成,每4个→3)-β-D-Glcp-(1→在O-6位连接一个长支链,该支链由α-D-Glcp-(1→、→4,6)-β-D-Glcp-(1→、→4)-β-D-Glcp-(1→和→6)-β-D-Glcp-(1→依次相连构成。结论通过膜技术和凝胶色谱分离纯化得到活性多糖肽,该方法简便、快速,为灵芝提取物及其产品中多糖肽质量控制提供了科学依据。

Objective To study the structure of active polysaccharide peptide purified from Ganoderma lucidum aqueous extract,and used as a reference substance for the determination of polysaccharide peptide content in G.lucidum products.Methods GL-PPSQ2 was obtained by hot water extraction,separation and purification with membrane ultrafiltration and gel-filtration chromatography.The physicochemical determination and spectral date were used for structural identification.The content of polysaccharide peptide was detected by HPLC with UV detector,water was used as mobile phase and the flow rate was 1 mL/min.Results GL-PPSQ2 was a pure polysaccharide peptide with purity above 97%,molecular weight of 5.0×10~4,polysaccharide content of 87.17%,and yield of 0.49%.The monosaccharides composition analysis showed that GL-PPSQ2 was glucose-based polysaccharide with a small amount of mannose,which contained 16 kinds of amino acids with the total amount of amino acids of 5.04%.Based on the methylation analysis,1D and 2D NMR spectroscopy,the repeating unit of GL-PPSQ2 was composed of →3)-β-D-Glcp-(1→backbone,with four repeating units connected a long chain branch at O-6 which was composed of α-D-Glcp-(1→,→4,6)-β-D-Glcp-(1→,→4)-β-D-Glcp-(1→ and →6)-β-D-Glcp-(1→ in sequence.Conclusion The active polysaccharide peptide was isolated and purified by membrane technology and gel chromatography.The method was simple and rapid,which provided a scientific basis for the quality control of polysaccharide peptide in G.lucidum extract and its products.