可口革囊星虫多糖酶法—碱梯次提取工艺优化及其抗氧化和抑菌性能分析
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摘要
【目的】优化可口革囊星虫多糖酶法—碱梯次提取工艺,并分析两阶段提取多糖的体外抗氧化和抑菌活性,为可口革囊星虫酶解产物及不溶性虫体成分的梯次化加工利用提供参考依据。【方法】以新鲜可口革囊星虫为材料、多糖提取率为考察指标,经木瓜蛋白酶和碱性蛋白酶处理,联合酶解和滤渣碱提工艺,两阶段梯次化提取多糖,并测定可口革囊星虫多糖对DPPH自由基和羟基自由基(·OH)的清除率,采用纸片法测定其抑菌性能。【结果】可口革囊星虫多糖酶法—碱梯次提取最佳工艺为:复合酶(木瓜蛋白酶∶碱性蛋白酶=1∶2)添加量4.0%、酶解温度55℃、酶解时间4 h、料液比1∶5(g/mL)、滤渣碱提时间2.0 h、氢氧化钠浓度1.5%,在此条件下,获得酶解多糖提取率为1.076%,滤渣碱提多糖提取率为0.171%,总多糖提取率为1.247%。酶提多糖和滤渣碱提多糖对·OH清除的半抑制浓度(IC50)分别为2.6和4.9 mg/mL,清除DPPH自由基的IC50分别为4.4和5.1 mg/mL。酶提多糖和碱提多糖在1.560 mg/mL以上对金黄色葡萄球菌、大肠埃希氏菌、铜绿假单胞菌和枯草芽孢杆菌均有一定的抑制作用。【结论】采用优化的酶法—碱梯次提取工艺可从可口革囊星虫酶解液和滤渣中有效提取多糖,且两种多糖均具有较强的抗氧化活性和抑菌作用。
【Objective】Extraction technology of enzymatic extraction of polysaccharides(EEP)and alkali extraction of polysaccharides from filter residues(AEPR)were studied by enzymolysis-alkali extraction from Phscolosoma esculenta. The antioxidant in vitro and antimicrobial activities of two-stage extraction of polysaccharides were tested. It provided reference for gradient processing of enzymatic hydrolysate and insoluble component of P. esculenta.【Method】Taking fresh P. esculenta as material,the extraction rate of polysaccharides was used as index while two-stage extraction of polysaccharides were extracted by enzymatic process of papain and alkaline protease combining with alkaline process of enzymatic hydrolysis residue. The DPPH free radical scavenging rate and hydroxyl(·OH)scavenging rate were detected,the antimicrobial properties were determined by paper disc method.【Result】The results showed that the optimum extraction process for enzymolysis-alkali echelon extraction of polysaccharides was:complex enzyme(papain∶alkaline protease=1∶2)adding amount was 4%,enzymolysis temperature was 55 ℃,enzymolysis time was 4 h,solid-liquid ratio was 1∶5(g/mL),residue alkaline extraction time was 2.0 h,concentration of alkali was 1.5%,the extraction rate of the EEP was1.076%,the extraction rate of the AEPR was 0.171%,and the total polysaccharide was 1.247%. Half-inhibitory concentration(IC50)of EEP and AEPR on·OH were 2.6 and 4.9 mg/mL,respectively,and the scavenging rate IC50 of DPPH free radical were 4.4 and 5.1 mg/mL. EEP and AEPR had certain inhibitory effects on Staphylococcus aureus,Escherichia coli,Pseudomonas aeruginosa and Bacillus subtilis at the concentration of more than 1.560 mg/mL.【Conclusion】The optimized enzymolysis-alkali echelon extraction of polysaccharides can extract polysaccharides from enzymatic hydrolysate and filter residue of P. esculenta,and the two polysaccharides have strong antioxidant activity and antimicrobial activity.
【Objective】Extraction technology of enzymatic extraction of polysaccharides(EEP)and alkali extraction of polysaccharides from filter residues(AEPR)were studied by enzymolysis-alkali extraction from Phscolosoma esculenta. The antioxidant in vitro and antimicrobial activities of two-stage extraction of polysaccharides were tested. It provided reference for gradient processing of enzymatic hydrolysate and insoluble component of P. esculenta.【Method】Taking fresh P. esculenta as material,the extraction rate of polysaccharides was used as index while two-stage extraction of polysaccharides were extracted by enzymatic process of papain and alkaline protease combining with alkaline process of enzymatic hydrolysis residue. The DPPH free radical scavenging rate and hydroxyl(·OH)scavenging rate were detected,the antimicrobial properties were determined by paper disc method.【Result】The results showed that the optimum extraction process for enzymolysis-alkali echelon extraction of polysaccharides was:complex enzyme(papain∶alkaline protease=1∶2)adding amount was 4%,enzymolysis temperature was 55 ℃,enzymolysis time was 4 h,solid-liquid ratio was 1∶5(g/mL),residue alkaline extraction time was 2.0 h,concentration of alkali was 1.5%,the extraction rate of the EEP was1.076%,the extraction rate of the AEPR was 0.171%,and the total polysaccharide was 1.247%. Half-inhibitory concentration(IC50)of EEP and AEPR on·OH were 2.6 and 4.9 mg/mL,respectively,and the scavenging rate IC50 of DPPH free radical were 4.4 and 5.1 mg/mL. EEP and AEPR had certain inhibitory effects on Staphylococcus aureus,Escherichia coli,Pseudomonas aeruginosa and Bacillus subtilis at the concentration of more than 1.560 mg/mL.【Conclusion】The optimized enzymolysis-alkali echelon extraction of polysaccharides can extract polysaccharides from enzymatic hydrolysate and filter residue of P. esculenta,and the two polysaccharides have strong antioxidant activity and antimicrobial activity.
引文
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董兰芳,张琴,童潼,许明珠.2015.方格星虫体腔液多糖的提取及体外抗氧化活性[J].食品研究与开发,36(11):46-49.[Dong L F,Zhang Q,Tong T,Xu M Z.2015.Extraction and antioxidant activity of polysaccharides from Sipunculus nudus coelomic fluid[J].Food Research and Development,36(11):46-49.]
黄岛平,刘永强,林葵,黄文琦,陈秋虹,陈建红,黄艳.2012.北部湾革囊星虫和方格星虫主要营养成分比较分析[J].大众科技,14(8):172-173.[Huang D P,Liu Y Q,Lin K,Huang W Q,Chen Q H,Chen J H,Huang Y.2012.Comparative analysis of main nutritive components on Phascolosoma esulenta and Sipunculus nudus in Beibu Gulf[J].Popular Science&Technology,14(8):172-173.]
孔帅.2014.可口革囊星虫多糖对小鼠免疫调节作用的研究[D].宁波:宁波大学.[Kong S.2014.Immunomodulatory effects of polysaccharides from Phascolosoma esulenta on mice[D].Ningbo:Ningbo University.]
孔帅,刘连亮,陆新江,史雨红,陈炯.2014.响应面法优化可口革囊星虫抗氧化多糖提取工艺研究[J].食品工业科技,35(11):233-237.[Kong S,Liu L L,Lu X J,Shi YH,Chen J.2014.Optimization of extraction process by response surface methodology of anti-oxidant polysaccharides from Phasolosma esculenta[J].Science and Technology of Food Industry,35(11):233-237.]
李凤鲁.1989.中国沿海革囊星虫属(星虫动物门)的研究[J].青岛海洋大学学报,19(3):78-90.[Li F L.1989.Studies on the genus Phascolosoma(Sipuncula)of the China coasts[J].Journal of Ocean University of Qingdao,19(3):78-90.]
李妍妍,徐巧云,郑卫星,丁理发,陈才利,苏秀榕.2007.可口革囊星虫体腔液多糖的研究[C]//中国药学会.2007年全国生化与生物技术药物学术年会论文集:527-528.[Li Y Y,Xu Q Y,Zheng W X,Ding L F,Chen C L,Su X R.2007.Studies on polysaccharides from Phasolosma esculenta coelomic fluid[C]//Chinese Pharmaceutical Association.Papers Collection of 2007 National Annual Conference on Biochemical and Biotechnical Pharmaceuticals:527-528.]
李永强.2011.北部湾(广西段)潮间带大型底栖动物的调查研究[D].青岛:青岛理工大学.[Li Y Q.2011.The macrozoobenthos of intertidal zone in Beibu Gulf(Guangxi)[D].Qingdao:Qingdao University of Technology.]
梁倩蓉,于曙光,刘顺,王国良,郑晓叶,彭雪美惠,周素明.2015.可口革囊星虫Phasolosma esculenta体腔液的抗菌活性研究[J].海洋学研究,33(2):70-75.[Liang Q R,Yu S G,Liu S,Wang G L,Zheng X Y,Peng X M H,Zhou S M.2015.Antibacterial activity of the coelomic fluid from a marine worm,Phasolosma esculenta[J].Journal of Marine Sciences,33(2):70-75.]
刘丽,徐春,李可,姜昆,吴鹏.2016.九资河茯苓多糖的超声波提取工艺及抑菌性研究(英文)[J].农业科学与技术,17(12):2746-2750.[Liu L,Xu C,Li K,Jiang K,Wu P.2016.Optimal ultrasonic extraction of pachyman from Jiuzihe Poria cocos[J].Agricultural Science&Technology,17(12):2746-2750.]
刘玉明,钱甜甜,何颖,蒋定文,沈先荣,江叔奇,杨进刚.2011.方格星虫多糖不同提取工艺的比较研究[J].时珍国医国药,22(6):1454-1455.[Liu Y M,Qian T T,He Y,Jiang D W,Shen X R,Jiang S Q,Yang J G.2011.Comparative study on polysaccharide from Sipunculus nudus by different extraction process[J].Lishizhen Medicine and Materia Medica Research,22(6):1454-1455.]
莫新.2017.可口革囊星虫高密度养殖技术[J].科学养鱼,(1):47.[Mo X.2017.High density breeding technology of the Phscolosoma esculenta[J].Scientific Fish Farming,(1):47.]
牛荣丽,唐健红.2012.可口革囊星虫抗疲劳作用的研究[J].食品工业科技,33(24):389-391.[Niu R L,Tang J H.2012.Research of anti-fatigue effect of Phascolosoma esculenta[J].Science and Technology of Food Industry,33(24):389-391.]
苏伟,宋玉,母应春,李泽秀,黎以栾.2017.响应面法优化木瓜蛋白酶提取红稗多糖工艺[J].食品工业科技,38(7):256-261.[Su W,Song Y,Mu Y C,Li Z X,Li Y L.2017.Optimization of extraction of polysaccharides from Baccage sedge using papain by response surface methodology[J].Science and Technology of Food Industry,38(7):256-261.]
田振华,李永斌,李颖,楚华航,郝丽丽,宿婷婷,张莉.2013.牡蛎多糖制备工艺优化及体外抑菌活性的探讨[J].食品研究与开发,34(17):23-26.[Tian Z H,Li Y B,Li Y,Chu H H,Hao L L,Su T T,Zhang L.2013.Optimization of preparation technology and antimicrobial activity of polysaccharide extracts from Rushan Bay oyster(Crassostrea gigas)in China[J].Food Research and Development,34(17):23-26.]
王冲.2005.蚯蚓(Eisenia fetida)生态免疫系统研究Ⅰ--体壁屏障系统及协同防御机理[D].北京:中国农业大学.[Wang C.2005.Research of ecology immunity system in the Eisenia fetidaⅠ-Antibiotic barrier system and mechanism of joint prevention[D].Beijing:Ocean University of China.]
王芳芳.2013.盐碱地生态养殖可口革囊星虫试验[J].科学养鱼,(5):73-74.[Wang F F.2013.A study on the ecological culture of the Phscolosoma esculenta in saline-alkali soil[J].Scientific Fish Farming,(5):73-74.]
王帅.2018.北部湾革囊星虫属资源初探及遗传多样性分析[D].南宁:广西大学.[Wang S.2018.The preliminary study on the resources of Phascolosoma in Beibu Gulf and its genetic diversity[D].Nanning:Guangxi University.]
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