海参硫酸软骨素的荧光标记方法

Fluorescence Labeling of Sea Cucumber Chondroitin Sulfate

海参硫酸软骨素是海参体壁的重要组成物质之一,已被证实具有多种生理调节功能,在功能食品开发方面展现出良好的应用前景。与其他碳水化合物类似,海参硫酸软骨素缺乏特异的发色及发光基团。基于此背景,本研究旨在对海参硫酸软骨素进行荧光标记。以6-氨基荧光素为荧光染料并配合溴化氢活化法成功对该多糖进行了荧光衍生,每分子标记产物中约含4.4分子荧光染料,其最佳荧光激发波长为495 nm、发射波长为520 nm。分子质量测定、硫酸根质量分数测定及傅里叶变换红外光谱比较分析的结果表明,标记前后多糖的结构特征未发生明显变化,标记产物在中性及碱性环境中展现出较强的荧光强度及稳定性,溶液环境中的NaCl对标记产物的影响有限。海参硫酸软骨素荧光标记方法的建立为后续研究该多糖的吸收代谢过程、功能机理、生物分子间相互作用等科学问题提供了有利条件。

Sea cucumber chondroitin sulfate(CHS)is one of the important bioactive substancesins in sea cucumber body wall.Since its multiple bioactive functions have been validated,CHS is considered as a potential ingredient of novel functional foods.Similar to most carbohydrates,CHS lacks specific chromophores and fluorescence groups.Based on this background,the aim of this study was to construct fluorescence labelling of CHS.The polysaccharide was derivatized by hydrogen bromide activation method with 6-amino-fluorescein as the fluorescent dye.As a result,each molecule of the labelled product approximately contained 4.4 molecules of 6-amino-fluorescein.The optimal fluorescence excitation and emission wavelengths of the labelled CHS were 495 and 520 nm,respectively.Additionally,its molecular weight and sulfate content were determined.Infrared spectroscopy showed that the structural characteristics of the polysaccharide did not significantly change after labeling.The labelled CHS exhibited strong fluorescence intensity and stability in neutral and alkaline environments.The influence of NaCl in the solution environment on the labelled CHS was limited.The establishment of this fluorescence labeling method will facilitate further research on the absorption and metabolism,functions and underlying mechanisms of CHS and its interactions with other biomolecules.