滑锈伞液体发酵培养基的优化及抗菌活性的初探
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摘要
以菌丝体生物量为指标,采用单因素试验及Plackett-Burman和Central composition design两种响应面法优化滑锈伞液体发酵培养条件;采用牛津杯法测定滑锈伞液体发酵液对金黄色葡萄球菌、枯草芽孢杆菌、绿脓假单胞菌和大肠杆菌的抗菌活性和最小抑菌浓度。结果表明,滑锈伞液体发酵的最优培养基为乳糖30. 26 g/L、牛肉膏3. 05 g/L、KCl 0. 5 g/L、MgSO_40. 5 g/L、K_2HPO_41 g/L和Fe SO_40. 01 g/L,菌丝生物量达(9. 15±0. 61)g/L,是初始发酵培养基的26. 60倍。滑锈伞液体发酵液对4种细菌的生长均有明显的抑制作用,其中对金黄色葡萄球菌抑制作用最强,抑菌圈直径为(24. 83±0. 39) mm,最低抑菌浓度为20 mg/mL。该文研究结果对于滑锈伞属真菌的高值利用具有一定的指导作用。
This study aimed to optimize the liquid cultivation medium for Hebeloma sp. and investigate its antibacterial activity. Single-factor experiment and Plackett-Burman and Central composition design response surface methodology were implemented by using the biomass of Hebeloma sp. as an indicator. Besides,the anti-bacterial activity and the minimal inhibitory concentration( MIC) of the medium against Escherichia coli,Staphyloccus aureus,Bacillus subtilis and Pesudomonas aeruginosa were investigated. It was found that the optimum fermentation medium consisted of 30. 26 g/L lactose,3. 05 g/L beef extract,0. 5 g/L KCl,0. 5 g/L MgSO_4,1 g/L K2 HPO_4,and 0. 01 g/L FeSO_4. The mycelial biomass of Hebeloma sp. reached( 9. 15 ± 0. 61) g/L under this condition,which was 26. 6 times higher than that under initial fermentation medium. Moreover,the medium had obvious effects on inhibiting the growths of E. coli,S. aureus,B. subtilis and P. aeruginosa. Besides,its inhibitory effect on S. aureus was the greatest,and the diameter of inhibitory zone was( 24. 83 ± 0. 39) mm and the MIC was 20 mg/mL. In conclusion,this study will be helpful for high-value utilization of Hebeloma sp.
This study aimed to optimize the liquid cultivation medium for Hebeloma sp. and investigate its antibacterial activity. Single-factor experiment and Plackett-Burman and Central composition design response surface methodology were implemented by using the biomass of Hebeloma sp. as an indicator. Besides,the anti-bacterial activity and the minimal inhibitory concentration( MIC) of the medium against Escherichia coli,Staphyloccus aureus,Bacillus subtilis and Pesudomonas aeruginosa were investigated. It was found that the optimum fermentation medium consisted of 30. 26 g/L lactose,3. 05 g/L beef extract,0. 5 g/L KCl,0. 5 g/L MgSO_4,1 g/L K2 HPO_4,and 0. 01 g/L FeSO_4. The mycelial biomass of Hebeloma sp. reached( 9. 15 ± 0. 61) g/L under this condition,which was 26. 6 times higher than that under initial fermentation medium. Moreover,the medium had obvious effects on inhibiting the growths of E. coli,S. aureus,B. subtilis and P. aeruginosa. Besides,its inhibitory effect on S. aureus was the greatest,and the diameter of inhibitory zone was( 24. 83 ± 0. 39) mm and the MIC was 20 mg/mL. In conclusion,this study will be helpful for high-value utilization of Hebeloma sp.
引文
[1]张敏,图力古尔.采自东北的中国滑锈伞属新纪录种[J].菌物学报,2017,36(8):1 168-1 175.
[2] MARMEISSE R,DEBAUD J C,CASSELTON L A. DNA probes for species and strain identification in the ectomycorrhizal fungus Hebeloma[J]. Mycological Research,1992,96(3):161-165.
[3] MAZUR X,BECKER U,ANKE T,et al. Two new bioactive diterpenes from Lepistasordida[J]. Phytochemistry,1996,43(2):405-407.
[4] KARDOSOVAA,MACHOVAE. Antioxidant activity of medicinal plant polysaccharides[J]. Fitoterapia,2006,77(5):367-373.
[5] WASSER S P. Medicinal mushrooms as a source of antitumor and immunomodulating polysaccharides[J]. Applied Microbiology and Biotechnology,2002,60(3):258-274.
[6]文庭池,李光荣,康冀川,等.蛹虫草液体种制备及发酵生产菌丝体和虫草菌素工艺优化[J].食品科学,2012,33(5):144-149.
[7]张晓娜,李云波,鲁云风,等.产紫杉醇内生真菌和食用菌液体发酵多糖含量比较研究[J].食品科技,2017,42(5):185-187.
[8]张婧迪,蔡明美,刘志诚,等.响应面设计优化绿僵菌固体发酵条件[J].微生物学通报,2016,43(9):2 072-2 078.
[9]吴遵秋,姜友军,苏光灿,等.油橄榄叶中橄榄苦苷的体外抗氧化和抑菌活性[J].食品科学,2014,35(21):94-99.
[10]冀瑞卿,李玉,宋瑞清.鹅膏菌产抑菌物质培养条件优化及安全性分析[J].东北林业大学学报,2012,40(3):98-102.
[11]任萌,高国富,咸漠,等.产α-蒎烯重组大肠杆菌发酵培养基优化[J].食品工业科技,2015,36(3):151-162.
[12]王维乐,牟志美,张淑君,等.响应面法优化Paraconiothyrium variabile GHJ-4产漆酶发酵条件[J].应用与环境生物学报,2015,17(3):321-325.
[13]张明亮,刘靖宇,常明昌,等.蛹虫草黄色素提取工艺优化及其体外抗氧化研究[J].菌物学报,2018,37(6):761-771.
[14]弓明钦,NICHOLAS MALAJCZUK,王凤珍.等.西澳粘滑菇在尾叶桉上的菌根合成[J].林业科学研究,1995,8(1):11-13.
[15] VUORINEN I,HAMBERG L,MULLER M,et al. Development of growth media for solid substrate propagation of ectomycorrhizal fungi for inoculation of Norway spruce(Piceaabies)seedlings[J]. Mycorrhiza,2015,25(4):311-324.
[16]图力古尔,包海鹰,李玉.中国毒蘑菇名录[J].菌物学报,2014,33(3):517-548.
[17] DOREJ,MARMEISSE R,COMBIER J P,et al. A fungal conserved gene from the basidiomycete Hebelomacylindrosporum is essential for efficient ectomycorrhiza formation[J]. Mol Plant Microbe Interact,2014,27(10):1 059-1 069.
[18] KORHONEN A, LEHTO T, HEINONEN J, et al.Whole-plant frost hardiness of mycorrhizal(Hebeloma sp.or Suillus luteus)and non-mycorrhizal Scots pine seedlings[J]. Tree Physiology,2018,https://doi. org/10.1093/treephys/tpy105.
[19] SIEMENS J A,CALVO-POLANCO M,ZWIAZEK J J.Hebelomacrustuliniforme facilitates ammonium and nitrate assimilation in trembling aspen(Populus tremuloides)seedlings[J]. Tree Physiology,2011,31(11):1 238-1 250.
[20] BECQUER A,GARCIA K,AMENC L,et al. The Hebelomacylindrosporum Hc PT2 Pi transporter plays a key role in ectomycorrhizal symbiosis[J]. New Phytol,2018,220(4):1 185-1 199.
[21]林陈强,严军,王衍霖,等.虎奶菇发酵液抑菌成分的理化性质及其分离鉴定[J].天然产物研究与开发,2014,26(4):534-538.
[22]李晨,张沛,肖林刚,等.金针菇不同极性部位的抑菌活性及稳定性[J].食品研究与开发,2017,38(2):29-33.
[23]周玉芝,齐玉臣,白育红.外生菌根真菌对植物根病原菌拮抗作用的研究[J].微生物学杂志,1991,11(1):14-18.
[24]邵红军,房立真,刘吉开.西澳粘滑菇的化学成分研究[J].天然产物研究与开发,2010,22(5):786-788.
[25] GARLASCHELLI L,VIDARI G,VIRTUANI M,et al.The structures of new lanostane triterpenes from the fruiting bodies of Hebelomasenescens[J]. Journal of Natural Products,1995,58(7):992-1 002.
[2] MARMEISSE R,DEBAUD J C,CASSELTON L A. DNA probes for species and strain identification in the ectomycorrhizal fungus Hebeloma[J]. Mycological Research,1992,96(3):161-165.
[3] MAZUR X,BECKER U,ANKE T,et al. Two new bioactive diterpenes from Lepistasordida[J]. Phytochemistry,1996,43(2):405-407.
[4] KARDOSOVAA,MACHOVAE. Antioxidant activity of medicinal plant polysaccharides[J]. Fitoterapia,2006,77(5):367-373.
[5] WASSER S P. Medicinal mushrooms as a source of antitumor and immunomodulating polysaccharides[J]. Applied Microbiology and Biotechnology,2002,60(3):258-274.
[6]文庭池,李光荣,康冀川,等.蛹虫草液体种制备及发酵生产菌丝体和虫草菌素工艺优化[J].食品科学,2012,33(5):144-149.
[7]张晓娜,李云波,鲁云风,等.产紫杉醇内生真菌和食用菌液体发酵多糖含量比较研究[J].食品科技,2017,42(5):185-187.
[8]张婧迪,蔡明美,刘志诚,等.响应面设计优化绿僵菌固体发酵条件[J].微生物学通报,2016,43(9):2 072-2 078.
[9]吴遵秋,姜友军,苏光灿,等.油橄榄叶中橄榄苦苷的体外抗氧化和抑菌活性[J].食品科学,2014,35(21):94-99.
[10]冀瑞卿,李玉,宋瑞清.鹅膏菌产抑菌物质培养条件优化及安全性分析[J].东北林业大学学报,2012,40(3):98-102.
[11]任萌,高国富,咸漠,等.产α-蒎烯重组大肠杆菌发酵培养基优化[J].食品工业科技,2015,36(3):151-162.
[12]王维乐,牟志美,张淑君,等.响应面法优化Paraconiothyrium variabile GHJ-4产漆酶发酵条件[J].应用与环境生物学报,2015,17(3):321-325.
[13]张明亮,刘靖宇,常明昌,等.蛹虫草黄色素提取工艺优化及其体外抗氧化研究[J].菌物学报,2018,37(6):761-771.
[14]弓明钦,NICHOLAS MALAJCZUK,王凤珍.等.西澳粘滑菇在尾叶桉上的菌根合成[J].林业科学研究,1995,8(1):11-13.
[15] VUORINEN I,HAMBERG L,MULLER M,et al. Development of growth media for solid substrate propagation of ectomycorrhizal fungi for inoculation of Norway spruce(Piceaabies)seedlings[J]. Mycorrhiza,2015,25(4):311-324.
[16]图力古尔,包海鹰,李玉.中国毒蘑菇名录[J].菌物学报,2014,33(3):517-548.
[17] DOREJ,MARMEISSE R,COMBIER J P,et al. A fungal conserved gene from the basidiomycete Hebelomacylindrosporum is essential for efficient ectomycorrhiza formation[J]. Mol Plant Microbe Interact,2014,27(10):1 059-1 069.
[18] KORHONEN A, LEHTO T, HEINONEN J, et al.Whole-plant frost hardiness of mycorrhizal(Hebeloma sp.or Suillus luteus)and non-mycorrhizal Scots pine seedlings[J]. Tree Physiology,2018,https://doi. org/10.1093/treephys/tpy105.
[19] SIEMENS J A,CALVO-POLANCO M,ZWIAZEK J J.Hebelomacrustuliniforme facilitates ammonium and nitrate assimilation in trembling aspen(Populus tremuloides)seedlings[J]. Tree Physiology,2011,31(11):1 238-1 250.
[20] BECQUER A,GARCIA K,AMENC L,et al. The Hebelomacylindrosporum Hc PT2 Pi transporter plays a key role in ectomycorrhizal symbiosis[J]. New Phytol,2018,220(4):1 185-1 199.
[21]林陈强,严军,王衍霖,等.虎奶菇发酵液抑菌成分的理化性质及其分离鉴定[J].天然产物研究与开发,2014,26(4):534-538.
[22]李晨,张沛,肖林刚,等.金针菇不同极性部位的抑菌活性及稳定性[J].食品研究与开发,2017,38(2):29-33.
[23]周玉芝,齐玉臣,白育红.外生菌根真菌对植物根病原菌拮抗作用的研究[J].微生物学杂志,1991,11(1):14-18.
[24]邵红军,房立真,刘吉开.西澳粘滑菇的化学成分研究[J].天然产物研究与开发,2010,22(5):786-788.
[25] GARLASCHELLI L,VIDARI G,VIRTUANI M,et al.The structures of new lanostane triterpenes from the fruiting bodies of Hebelomasenescens[J]. Journal of Natural Products,1995,58(7):992-1 002.