氯化胆碱/乳酸/愈创木酚三元共熔溶剂提取灵芝多糖的工艺及其抗氧化活性研究

Ganoderma lucidum Polysaccharides Extraction by Choline Chloride/Lactic Acid/Guaiacol Ternary Deep Eutectic Solvent and Its Antioxidant Activities

本研究采用氯化胆碱/乳酸/愈创木酚三元共熔溶剂(DES)来提取灵芝多糖(GLPs),优化DES溶剂提取灵芝多糖的工艺参数,采用傅里叶变换红外光谱(Fourier Transform Infrared Spectroscopy,FT-IR)和凝胶渗透色谱(Gel Permeation Chromatography,GPC)对GLPs结构进行了表征,并研究其体外抗氧化活性,最后,采用密度泛函理论初步研究了DES提取GLPs的分子机制。结果表明,在含水量为50%、提取温度为80℃、提取时间为1 h、料液质量体积比为1:30 g/mL的条件下,GLPs得率达9.20%±0.27%。FT-IR和GPC分析表明,4000~500 cm-1区域GLPs具有典型的多糖结构,GLPs的数均分子量为8057 Da,多分散系数为1.6。体外抗氧化活性测试显示,当GLPs质量浓度为5 mg/mL时,其总还原力为0.67±0.04。此外,GLPs对DPPH、OH和ABTS+自由基的清除能力的IC50值分别为2.366、6.179和2.440 mg/mL。密度泛函理论模拟分析表明,DES与葡萄糖之间的结合能为65.29 kcal/mol,远高于水与葡萄糖之间的结合能(20.38 kcal/mol),DES提取多糖是通过三重氢键相互作用来实现的。研究成果从分子水平上阐明了DES提取GLPs效果优于传统热水提取方法的内在机制。

In this study,choline chloride/lactic acid/guaiacol ternary deep eutectic solvent(DES)was used to extract Ganoderma lucidum polysaccharides(GLPs).The operational parameters of GLPs extraction by DES were optimized,and the obtained GLPs structure was characterized by Fourier transform infrared spectrophotometer(FT-IR)and gel chromatography(GPC).Furthermore,the in vitro antioxidant activities of GLPs were investigated.In the end,density functional theory(DFT)was leveraged to gain preliminary insights into the molecular mechanism underpinning polysaccharide extraction facilitated by DES.Results unveiled that when the moisture content within DES was maintained at 50%,extraction was carried out at a temperature of 80℃for 1 h,and the liquid-to-solid mass-volume ratio stood at 1:30 g/mL,the extraction yield of GLPs reached 9.20%±0.27%.FT-IR and GPC results showed that GLPs had typical polysaccharide structures in the 4000~500 cm-1 regions,and the number average mass weight(Mn)of GLPs was 8057 Da with the polydispersity index(PDI)of 1.6.In the context of in vitro antioxidant activity experiments,when GLPs content was 5 mg/mL,the total reducing power was measured at 0.67±0.04.Notably,the IC50 values for GLPs'scavenging capabilities against DPPH,OH,and ABTS+free radicals were determined to be 2.366,6.179,and 2.440 mg/mL,respectively.DES simulation analyses revealed a binding energy of 65.29 kcal/mol between DES and glucose,a substantially higher value compared to the binding energy observed between water and glucose(20.38 kcal/mol).The GLPs extraction by DES was facilitated through the pivotal role of triple hydrogen bond interactions.The findings in this study elucidated the intrinsic mechanism at molecular level that underscores the superior effectiveness of DES in comparison to traditional hot water extraction methods.