金钗石斛多糖的酶解工艺优化及其抗氧化活性  

Process optimization of Dendrobium nobile polysaccharide on enzymatic hydrolysis and its antioxidant activity

作　　者：王筱玥 刘学 刘春环 刘晶 杨成 WANG Xiaoyue;LIU Xue;LIU Chunhuan;LIU Jing;YANG Cheng(School of Chemical and Material Engineering,Jiangnan University,Wuxi 214122,Jiangsu,China;Guozhen Health Technology(Beijing)Co.,Ltd.,Beijing 102200,China)

机构地区：[1]江南大学化学与材料工程学院,江苏无锡214122 [2]国珍健康科技(北京)有限公司,北京102200

出　　处：《精细化工》2025年第4期837-848,共12页Fine Chemicals

摘　　要：采用α-半乳糖苷酶对金钗石斛多糖30%醇沉(30%为乙醇水溶液中乙醇的质量分数)组分(DNP-30)进行了酶解。以酶解产物(DNP-30E)对2,2-联氮-二(3-乙基-苯并噻唑-6-磺酸)二铵盐(ABTS+)自由基清除率为指标,通过单因素和响应面实验确定了最佳酶解条件。采用FTIR、NMR、高效凝胶渗透色谱仪对DNP-30和DNP-30E进行了表征。基于体外和细胞内抗氧化活性实验、建立十二烷基硫酸钠(SDS)刺激HaCaT细胞氧化应激模型,探究了DNP-30和DNP-30E的抗氧化能力。结果表明,在α-半乳糖苷酶加酶量20 U/mL、pH=7.0、酶解温度46℃、酶解时间11 h的最佳酶解条件下,DNP-30E对ABTS+自由基清除率平均值为85.62%。与DNP-30相比,DNP-30E还原糖含量由0.63%±0.35%升至48.66%±0.51%,重均相对分子质量由7.95×10^(5)Da降至2083 Da。酶解过程造成多糖中的葡萄糖(1α→6)-甘露糖〔Glu(1α→6)-Man〕键发生断裂,并引起葡萄糖端基氢构型翻转。当多糖质量浓度为2.0 g/L时,DNP-30E提高了ABTS+自由基、羟基自由基、超氧自由基清除率;当多糖质量浓度为5.0~10.0 g/L时,DNP-30E细胞毒性比DNP-30显著降低。多糖质量浓度为0.5 g/L时,DNP-30E组比DNP-30组的活性氧含量降低了34.21%,丙二醛含量降低了50.2%,超氧化物歧化酶活力提升了84.17%,表现出更强的抗氧化能力。α-Galactosidase was used to enzymolyze 30%alcohol precipitation(30%is the mass fraction of ethanol in ethanol aqueous solution)component(DNP-30)of Dendrobium nobium polysaccharide,and the optimal enzymolysis conditions were determined by single factor and response surface experiments using scavenging rate of 2,2-diazo-bis(3-ethyl-benzothiazole-6-sulfonic acid)diamiammonium salt(ABTS+)free radicals by enzymolysis product(DNP-30E)as index.DNP-30 and DNP-30E were characterized by FTIR,NMR and high performance gel permeation chromatograph.The oxidative stress model of HaCaT cells stimulated by sodium dodecyl sulfate(SDS)was established to investigate the antioxidant capacity of DNP-30 and DNP-30E based on in vitro and intracellular antioxidant activity experiments.The results showed that under the optimum enzymolysis conditions ofα-galactosidase dosage 20 U/mL,pH=7.0,temperature 46℃and time 11 h,the average scavenging rate of DNP-30E on ABTS+free radicals was 85.62%.Compared with those of DNP-30,the reducing sugar content of DNP-30E increased from 0.63%±0.35%to 48.66%±0.51%,and the mass-average relative molecular mass decreased from 7.95×10^(5)Da to 2083 Da.Enzymolysis might break glucose(1α→6)-mannose[Glu(1α→6)-Man]bond in DNP-30 and reversed the configuration of glucose terminal hydrogen.When the mass concentration of polysaccharide was 2.0 g/L,DNP-30E increased the scavenging rate of ABTS+free radicals,hydroxyl free radicals,superoxide free radicals.When the mass concentration of polysaccharide was 5.0~10.0 g/L,the cytotoxicity of DNP-30E was significantly lower than that of DNP-30.When polysaccharide concentration was 0.5 g/L,the active oxygen content,malondialdehyde content and superoxide dismutase activity of DNP-30E group were decreased by 34.21%,50.2%and 84.17%,respectively,compared with DNP-30 group,showing stronger antioxidant capacity.

关 键 词：金钗石斛多糖 Α-半乳糖苷酶 酶解工艺优化 抗氧化活性 HACAT细胞 中药现代化技术 

分 类 号：R284[医药卫生—中药学]