发酵虫草菌丝体多糖提取条件优化及其结构分析
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摘要
采用响应面法优化蝙蝠蛾拟青霉菌丝体多糖的提取工艺,并对提取所得多糖的结构进行初步分析。结果表明:发酵虫草菌丝体多糖提取最佳工艺条件为提取温度98.0℃、提取时间4.5 h、液固比25∶1(mL/g),该条件下多糖得率为9.69%。菌丝体多糖主要由中性糖(84.7%)和糖醛酸(9.5%)组成。采用高效体积排阻色谱、红外光谱扫描和离子色谱对多糖结构进行光谱分析和单糖组成分析,结果表明该多糖主要为分子质量61.6 kD的单一色谱峰且可能呈吡喃环结构。另外,菌丝体多糖由阿拉伯糖、半乳糖、葡萄糖、木糖、甘露糖和半乳糖醛酸组成,其物质的量比为2.5∶31∶36∶1∶15∶4。研究成果可为日后发酵虫草菌丝体多糖精细结构与活性研究以及产品开发提供一定理论依据。
Response surface methodology was applied to optimize polysaccharide extraction from Paecilomyces hepiali Chen & Dai. The extracted polysaccharide was structurally elucidated. The optimal extraction conditions were found to be 4.5 h extraction with hot water at 98.0 ℃ at water/material ratio of 25:1(m L/g). Under these conditions, the maximum polysaccharide yield of 9.69% was obtained and the polysaccharide mainly composed of neutral polysaccharide(84.7%) and uronic acid(9.5%). High performance size exclusion chromatography(HPESC), high performance anion exchange chromatography and FT-IR spectroscopy were used to analyze the structural characteristics of the polysaccharide. Results showed that the molecular weight of the polysaccharide was 61.6 kD, displaying a single chromatographic peak, and its structure contained a pyran ring. In addition, the polysaccharide was composed of arabinose, galactose, glucose, xylose, mannose and galacturonic acid, with a molar ratio of 2.5:31:36:1:15:4. The findings may be useful for future studies on the structure-activity relationship of polysaccharides from Paecilomyces hepiali Chen & Dai and for the development of new polysaccharide-based products.
Response surface methodology was applied to optimize polysaccharide extraction from Paecilomyces hepiali Chen & Dai. The extracted polysaccharide was structurally elucidated. The optimal extraction conditions were found to be 4.5 h extraction with hot water at 98.0 ℃ at water/material ratio of 25:1(m L/g). Under these conditions, the maximum polysaccharide yield of 9.69% was obtained and the polysaccharide mainly composed of neutral polysaccharide(84.7%) and uronic acid(9.5%). High performance size exclusion chromatography(HPESC), high performance anion exchange chromatography and FT-IR spectroscopy were used to analyze the structural characteristics of the polysaccharide. Results showed that the molecular weight of the polysaccharide was 61.6 kD, displaying a single chromatographic peak, and its structure contained a pyran ring. In addition, the polysaccharide was composed of arabinose, galactose, glucose, xylose, mannose and galacturonic acid, with a molar ratio of 2.5:31:36:1:15:4. The findings may be useful for future studies on the structure-activity relationship of polysaccharides from Paecilomyces hepiali Chen & Dai and for the development of new polysaccharide-based products.
引文
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[27]许峰,吴玲芳,林善,等.发酵冬虫夏草菌丝体中虫草多糖含量的检测及结构鉴定[J].基因组学与应用生物学,2014(6):1294-1302.
[28]黄静涵,艾斯卡尔·艾拉提,毛健.灵芝多糖的分离纯化及结构鉴定[J].食品科学,2011,32(12):301-304.
[29]ZHOU X,XU L,FENG S,et al.Characteristics and bioactivities of different molecular weight polysaccharides from camellia seed cake[J].International Journal of Biological Macromolecules,2016,91:1025-1032.DOI:10.1016/j.ijbiomac.2016.06.067.
[30]WU F,YAN H,MA X,et al.Comparison of the structural characterization and biological activity of acidic polysaccharides from Cordyceps militaris,cultured with different media[J].World Journal of Microbiology&Biotechnology,2012,28(5):2029-2038.DOI:10.1007/s11274-012-1005-6.
[2]LEE S,LEE H H,KIM J,et al.Anti-tumor effect of Cordyceps militaris,in HCV-infected human hepatocarcinoma 7.5 cells[J].Journal of Microbiology,2015,53(7):468-474.DOI:10.1007/s12275-015-5198-x.
[3]王林萍,余意,冯成强.冬虫夏草活性成分及药理作用研究进展[J].中国中医药信息杂志,2014(7):132-136.DOI:10.3969/j.issn.1005-5304.2014.07.043.
[4]刘高强,王晓玲,杨青,等.冬虫夏草化学成分及其药理活性的研究[J].食品科技,2007,32(1):202-205.DOI:10.3969/j.issn.1005-9989.2007.01.061.
[5]张姝,张永杰,SHRESTHA B,等.冬虫夏草菌和蛹虫草菌的研究现状、问题及展望[J].菌物学报,2013,32(4):577-597.
[6]丘雪红,曹莉,韩日畴.冬虫夏草的研究进展、现存问题与研究展望[J].环境昆虫学报,2016,38(1):1-23.DOI:10.3969/j.issn.1674-0858.2016.01.1.
[7]詹小涛,赖桂萍,朱德霞.冬虫夏草及其发酵菌丝体药理作用研究进展[J].医学综述,2012,18(10):1566-1569.
[8]曾世明.蝙蝠蛾拟青霉多糖的结构及活性研究[D].长春:东北师范大学,2011.
[9]WANG J,LI L Z,LIU Y G,et al.Investigations on the antifatigue and antihypoxic effects of Paecilomyces hepiali extract[J].Molecular Medicine Reports,2016,13(2):1861-1868.DOI:10.3892/mmr.2015.4734.
[10]WU Z,YANG Z,DAN G,et al.Influences of carbon sources on the biomass,production and compositions of exopolysaccharides from Paecilomyces hepiali,HN1[J].Biomass&Bioenergy,2014,67:260-269.DOI:10.1016/j.biombioe.2014.05.008.
[11]钟石,李有贵,陈诗,等.人工培养蛹虫草与冬虫夏草的主要活性成分比较[J].蚕业科学,2009,35(4):831-836.DOI:10.3969/j.issn.0257-4799.2009.04.020.
[12]WANG J,KAN L,NIE S,et al.A comparison of chemical composition,bioactive components and antioxidant activity of natural and cultured Cordyceps sinensis[J].Lebensmittel-Wissenschaft undTechnologie,2015,63(1):2-7.DOI:10.1016/j.lwt.2015.03.109.
[13]DENG B,WANG Z P,TAO W J,et al.Effects of polysaccharides from mycelia of Cordyceps sinensis,on growth performance,immunity and antioxidant indicators of the white shrimp Litopenaeus vannamei[J].Aquaculture Nutrition,2014,21(2):173-179.DOI:10.1111/anu.12147.
[14]WANG Y,WANG M,LING Y,et al.Structural determination and antioxidant activity of a polysaccharide from the fruiting bodies of cultured Cordyceps sinensis[J].American Journal of Chinese Medicine,2009,37(5):977-989.
[15]ZHANG J,YU Y,ZHANG Z,et al.Effect of polysaccharide from cultured Cordyceps sinensis on immune function and antioxidation activity of mice exposed to 60Co[J].International Immunopharmacology,2011,11(12):2251-2257.DOI:10.1016/j.intimp.2011.09.019.
[16]李景恩,聂少平,杨超,等.响应曲面法优化香薷多糖的提取工艺[J].食品科学,2009,30(18):131-134.DOI:10.3321/j.issn:1002-6630.2009.18.026.
[17]ZHANG H,NIE S P,YIN J Y,et al.Structural characterization of a heterogalactan purified from fruiting bodies of Ganoderma atrum[J].Food Hydrocolloids,2014,36(5):339-347.DOI:10.1016/j.foodhyd.2013.08.029.
[18]陈雪峰,贾士儒,王岳,等.发菜多糖的红外光谱分析与抗氧化活性的研究[J].食品与发酵工业,2009(7):133-137.
[19]LI W,WANG Q,CUI S W,et al.Elimination of aggregates of(1→3)(1→4)-β-D-glucan in dilute solutions for light scattering and size exclusion chromatography study[J].Food Hydrocolloids,2006,20(2):361-368.DOI:10.1016/j.foodhyd.2005.03.018.
[20]王海燕,戴军,陈尚卫.灵芝菌丝体多糖的分离纯化及其单糖组成分析与分子质量测定[J].食品与机械,2015(5):201-205.
[21]PU X,MA X,LU L,et al.Structural characterization and antioxidant activity in vitro of polysaccharides from angelica,and astragalus[J].Carbohydrate Polymers,2016,137:154-164.DOI:10.1016/j.carbpol.2015.10.053.
[22]张攀峰,陈玲,李晓玺,等.不同直链/支链比的玉米淀粉分子质量及其构象[J].食品科学,2010,31(19):157-160.
[23]康继.天然高性能乳化剂—印度树胶的精细分子结构和构象特性研究[D].无锡:江南大学,2011.
[24]杨涛,郭龙,李灿,等.红芪多糖HPS1-D的化学结构和抗补体活性研究[J].中国中药杂志,2014,39(1):89-93.DOI:10.4268/cjcmm20140118.
[25]WU Zhongwei,ZHANG Mingxia,XIE Minhao,et al.Extraction,characterization and antioxidant activity of mycelial polysaccharides from Paecilomyces hepiali HN1[J].Carbohydrate Polymers,2015,137(3):263-298.DOI:10.1016/j.carbpol.2015.11.010.
[26]刘金花,李富奎,贾得儒,等.中国被毛孢发酵虫草菌丝体多糖的提取、纯化及其理化性质[J].食品与发酵工业,2014,40(3):222-226.
[27]许峰,吴玲芳,林善,等.发酵冬虫夏草菌丝体中虫草多糖含量的检测及结构鉴定[J].基因组学与应用生物学,2014(6):1294-1302.
[28]黄静涵,艾斯卡尔·艾拉提,毛健.灵芝多糖的分离纯化及结构鉴定[J].食品科学,2011,32(12):301-304.
[29]ZHOU X,XU L,FENG S,et al.Characteristics and bioactivities of different molecular weight polysaccharides from camellia seed cake[J].International Journal of Biological Macromolecules,2016,91:1025-1032.DOI:10.1016/j.ijbiomac.2016.06.067.
[30]WU F,YAN H,MA X,et al.Comparison of the structural characterization and biological activity of acidic polysaccharides from Cordyceps militaris,cultured with different media[J].World Journal of Microbiology&Biotechnology,2012,28(5):2029-2038.DOI:10.1007/s11274-012-1005-6.