超声波降解改善黑穗醋栗果实多糖降血糖活性作用机制及构效关系研究

Aim at lack of hypoglycemic drugs to safely and effectively control diabetes, sonication, hypoglycemic activity as well as polysaccharides structure are combined to investigate. Mechanism of ultrasonic degradation improving the hypoglycemic activity of polysaccharides from black currant fruit and the structure-activity relationships are studied. Firstly, series of different molecular weight polysaccharides were prepared by ultrasonic degradation and the ultrasound degradation pathway and basic law of polysaccharides was studied. Secondly, the combination of carbohydrate chemistry, spectroscopy and chromatography methods was used to study the effect of ultrasound degradation on the primary structure and conformation of polysaccharides from black currant fruit. In addition, cell culture techniques, Real-time RT-PCR and Western blot were used to study the effect of ultrasonic degradation on the hypoglycemic activity of polysaccharides. On these researches basis, green ultrasonic modification technology of polysaccharides was established, the mechanism of ultrasonic degradation improving the hypoglycemic activity of polysaccharides and the structure-activity relationship were clarified, and the primary structure and conformation of plant polysaccharides with higher hypoglycemic activity was determined. This project provide an important theoretical support and technical guidance for the molecular design and recombinant for the future development of black currant fruit polysaccharide hypoglycemic agents. It is significant for the diabetes prevention.

针对安全、有效控制糖尿病的降血糖药物缺乏的问题，本项目将超声波降解、降血糖活性和多糖结构三者有机结合，研究超声波降解改善黑穗醋栗果实多糖降血糖活性作用机制以及多糖结构与降血糖活性之间的构效关系。首先采用超声波降解制备系列分子量均一多糖，研究降解途径与降解规律；再通过糖化学、光谱学和色谱学等学科相结合的方法研究超声波降解对黑穗醋栗果实多糖一级结构与空间构象的影响；采用细胞培养技术、Real-time RT-PCR、Western blot等手段系统研究超声波降解前后黑穗醋栗果实均一多糖的降血糖活性。通过上述研究，建立多糖的绿色环保超声波改性技术，探明超声波降解改善黑穗醋栗果实均一多糖降血糖活性的作用机制及其构效关系，明确具有较高降血糖活性的植物多糖的一级结构与空间构象。本项目为植物多糖降血糖药物的研制而进行分子设计和重组提供重要的理论支持和技术指导，对于糖尿病的防治具有重要意义。

目前，如何对糖尿病患者进行合理友好的治疗和日常保健，特别是从天然植物中寻找和开发安全、有效、方便使用的大众化降血糖产品，已成为国内外医药、化学工作者所关注的重要课题。本研究将超声波降解、降血糖活性和多糖结构三者有机结合，首先采用加压水法提取得到黑穗醋栗果实粗多糖，经柱色谱分离纯化后得到均一多糖；以黑穗醋栗果实高分子量均一多糖为原料，制备不同超声处理条件(时间、功率)下多糖样品，共计36种多糖。通过研究超声波参量对黑穗醋栗果实均一多糖分子量及分布、结构特性的影响，系统分析黑穗醋栗果实均一多糖在超声波作用下的降解规律，确定超声降解多糖的动力学模型。再通过糖化学、光谱学和色谱学等学科相结合的方法研究超声波降解对黑穗醋栗果实多糖一级结构和空间构象以及降血糖活性的影响，初步探讨超声波降解改善黑穗醋栗果实多糖降血糖活性作用机制，筛选出降血糖活性较高的黑穗醋栗果实多糖组分。结果表明：在三种超声功率分别作用下，多糖的降解规律均符合二级反应动力学模型（R2 > 0.98）及中点断链模型（R2 > 0.98）；超声波处理能够提高多糖的热稳定性，降低多糖溶液的表观黏度，削弱溶液的剪切稀化现象；超声波不会改变多糖的官能团、单糖残基的种类及糖苷键的类型；扫描电镜及原子力显微镜观测结果表明：伴随超声功率增大，降解多糖的片状面积逐渐减小，聚集度逐渐降低；随着超声功率增大，得到降解多糖的降血糖活性也逐步提高，且降解多糖对α-淀粉酶、α-葡萄糖苷酶抑制类型为可逆性竞争抑制；初步确定具有较高降血糖活性多糖U-700的一级结构与空间构象，并初步推断出其多糖链的精细结构。本项目为建立多糖的绿色环保的超声波改性技术奠定了理论基础，也为改善小浆果及其他植物多糖降血糖活性等相关研究提供新的思路。