响应面法优化龙须菜多糖快速溶剂萃取提取工艺及其抗炎活性研究

Optimization of Accelerated Solvent Extraction of Polysaccharides from Gracilaria lemaneiformis Using Response Surface Methodology and Anti-inflammatory Activity

为了建立快速溶剂萃取技术(ASE)提取龙须菜多糖的新方法,本文以秦皇岛龙须菜为原料,利用ASE提取龙须菜粗多糖(GLP-K),以多糖得率为指标,采用单因素实验结合响应面试验法优化提取工艺条件。通过傅里叶红外光谱和高效液相色谱对多糖进行结构表征;探索GLP-K在脂多糖(LPS)诱导的RAW264.7巨噬细胞中的抗炎作用。结果表明,快速溶剂萃取法用于龙须菜多糖提取的最佳工艺参数为提取温度70℃,提取时间8.5 min,循环4次,在此条件下,多糖实验得率为9.58%±0.31%;红外光谱证实该多糖含有糖醛酸,重均分子量在4.4~747.1 kDa之间;GLP-K在浓度1000μg/mL及以下时对RAW264.7细胞增殖也无影响(P<0.001);与模型组相比,GLPK给药组(50、100、200、300、400、500μg/mL)NO的释放量显著降低43.76%~69.47%(P<0.001)。本文丰富了快速溶剂萃取技术在多糖提取方面的研究,为龙须菜多糖的开发和利用提供实验依据。

The aim of this work was to apply response surface methodology(RSM)to model and optimize the accelerated solvent extraction(ASE)technique for extracting the crude polysaccharides(GLP-K)from Gracilaria lemaneiformis.In terms of the yield of polysaccharide,single factor tests and Box-Behnken design response surface method were developed.The structure of the prepared polysaccharides was characterized by Fourier transform infrared(FT-IR)and high performance liquid chromatography(HPLC).And the anti-inflammatory potential of GLP-K in lipopolysaccharide(LPS)-induced RAW264.7 macrophages was also explored.Briefly,the optimal extraction conditions for GLP-K were as follows:70°C of extraction temperature,8.5 min of extraction time,and 4 extraction cycles.Under the condition,the experimental yield of polysaccharide was 9.58%±0.31%.FT-IR showed that the polysaccharide contained uronic acid,and the weightaverage molecular weight ranged from 4.4 to 747.1 kDa.GLP-K had no significant cytotoxic effects at or below the concentrations of 1000μg/mL(P<0.001).Compared with the model group,GLP-K administration group(50,100,200,300,400,500μg/mL)displayed remarkable inhibitory effects on the release level of NO(P<0.001),which decreased by 43.76%~69.47%.This paper enriched the research on the extraction of polysaccharides by accelerated solvent extraction technology,and provided experimental basis for the development and utilization of Gracilaria lemaneiformis polysaccharides.