摘要
“桑黄”[曾被广泛称为Phellinus linteus (Berk.&M.A. Curtis) Teng]是我国一类传统药用真菌的俗称,但一直以来,该类群真菌的分类与系统发育研究一直存在争议和不明确性。目前,称为“桑黄”的药用真菌的研究和产品开发如火如荼,但同时存在的名称混乱现象也日益严重,极大地制约了这一宝贵药用真菌资源的科学利用和健康可持续发展。为此,本研究通过对来自东亚、美洲和非洲大量标本的研究,明确了该类群种类的形态分类依据和系统发育关系以及与地理分布和寄主间的联系。另外,首次对桑黄纤孔菌Inonotus sanghuang Sheng H. Wu, T. Hatt.&Y.C. Dai进行了液体发酵以及有效活性成分和体外抗氧化活性的研究。
桑黄类群(Inonotus linteus complex)包括多个形态特征类似的种：子实体盖形,菌肉菌丝一体系,菌管菌丝二体系的多样的菌丝系统。子实层具刚毛,担孢子宽椭圆形至近球形,黄色,厚壁。本论文研究中,将来自中美洲的一个裂蹄纤孔菌Inonotus linteus标本,作为该种的研究材料,完善并进一步界定了狭义上该种的形态和系统发育学信息。据此,来自中美洲的一个新种：古巴纤孔菌I. cubensis Y.C. Dai, Decock&L.W. Zhou和来自东亚地区的两个新种：高山纤孔菌I. alpinus Y.C. Dai&X.M. Tian和环区纤孔菌I. zonatus Y.C. Dai&X.M. Tian被发现。基于ITS序列进行的系统发育学分析也进一步明确了该类群中其它8个种：鲍姆纤孔菌I. baumii T. Wagner&M. Fisch.,裂蹄纤孔菌I. linteus (Berk.&M.A. Curtis) Teixeira,小孔忍冬纤孔菌lonicericola (Parmasto) Y.C. Dai,大孔忍冬纤孔菌I. lonicerinus (Bondartsev) Sheng H. Wu,Y.C. Dai&T. Hatt.,瓦宁纤孔菌I. vaninii (Ljub.) T. Wagner&M. Fisch.,桑黄纤孔菌I. sanghuang Sheng H. Wu, T. Hatt.&Y.C. Dai,锦带花纤孔菌I. weigelae T. Hatt.&Sheng H. Wu和韦尔纤孔菌I. weirianus (Bres.) T. Wagner&M. Fisch在该类群中的独立分枝地位。I. tenuicontextus L.W. Zhou&W.M. Qin经研究证实是I. weigelae的同物异名。另外,还给出了一个关于该类群11个种的形态分类检索表。
对于该类群中专性寄生于桑树上的桑黄纤孔菌I. sanghuang,利用响应面分析的方法进行了液体发酵培养基的优化。首先应用N=12的Plackett-Burman设计法对影响桑黄纤孔菌菌丝生物量的发酵培养基组分进行筛选,选取的8个相关因子为：玉米粉、葡萄糖、蛋白胨、酵母抽提物、麸皮、KH2PO4、MgSO47H2O和发酵液起始pH。统计分析表明,影响桑黄纤孔菌菌丝生物量的关键因子为玉米粉、酵母抽提物、麸皮和pH。在进行最陡爬坡试验逼近4个关键因素的最大响应区域的基础上,采用4因素3水平的Box-Behnken设计法对发酵培养基组分进行优化,获得的最佳培养条件为：玉米粉38.96g/L,葡萄糖25.00g/L,蛋白胨3.75g/L,酵母抽提物4.15g/L,麸皮20.55g/L, KH2PO41.25g/L, MgSO47H2O0.625g/L和pH6.39。在此条件下,桑黄纤孔菌菌丝生物量预测值和验证值分别为17.60g/L和18.33±0.86g/L,达优化前(7.29g/L)的2.5倍。
利用水提醇沉法提取得到桑黄纤孔菌液体发酵胞内多糖(IPS)和胞外多糖(EPS)粗提物后,通过葡聚糖凝胶Sephadex G-100进行分级纯化,分别获得两个胞内多糖组分IPS1、IPS2和两个胞外多糖组分EPS1、EPS2。通过分析6个桑黄纤孔菌多糖样品的多糖含量和体外清除DPPH自由基活性的结果发现,供试6个多糖样品的多糖含量与体外清除DPPH自由基活性没有明显的相关性；桑黄纤孔菌胞内多糖的3个样品在6.25μg/mL至500μg/mL浓度范围内,DPPH自由基清除率随样品浓度增高而逐渐增强,具有一定的体外抗氧化活性；桑黄纤孔菌胞外多糖的3个样品在试验浓度范围内没有明显的清除DPPH自由基活性。
为探讨两阶段培养模式对桑黄纤孔菌液体发酵代谢产物积累的影响,在振荡培养10d后,继续静置培养30d,并跟踪测定了桑黄纤孔菌菌丝体中总黄酮、总多酚和总多糖的含量变化以及菌丝体醇提物(IS-EtOH)和水提物(IS-H2O)体外清除DPPH自由基和羟自由基的活性变化情况。试验结果显示：菌丝体中总黄酮和总多酚的含量在经过10d和15d的静置培养后,分别提高了37.92%和77.27%；随静置培养时间的延长,菌丝体醇提物中黄酮类组分的种类不断丰富,根据初步的HPLC检测结果,推测可能含有：芦丁、柚皮素、圣草酚和樱花亭4种黄酮类成分；桑黄纤孔菌菌丝体中酚类结构的代谢产物(总黄酮和总多酚)在桑黄纤孔菌菌丝体提取物体外清除DPPH自由基和羟自由基方面,发挥了主要作用。
"Sanghuang" is a popular medicinal mushroom complex in traditional Chinese medicine, and was previously recorded as Phellinus linteus (Berk.&M.A. Curtis) Teng. But for a long time, the taxonomy and phylogeny of this complex has been controversial and ambiguous. Currently studies and products on medicinal mushroom called "sanghuang" in full swing, but at the same time the phenomenon of species confusion is increasingly. It has greatly restricted the scientific exploitation, healthy and sustainable development of this precious medicinal mushroom. So the research on a plenty of specimens from East Asia, American and Africa was carried out to determine the morphological and phylogenetic relationships and their geographical distribution and hosts. In addition, the study about liquid fermentation, bioactive components and antioxidant activity in vitro were conducted on Inonotus sanghuang Sheng H. Wu, T. Hatt.&Y.C. Dai.
The Inonotus linteus complex comprised several closely related species characterized by pileate basidiomes, a heterogeneous hyphal system with a monomitic context and a dimitic hymenophoral trama. Setae are from hymenia and basidiospores are broadly ellipsoid to subglobose, yellowish and thick-walled. Researches to understand this complex have been particularly notable in East Asia. In the present paper, the complex was studied at a larger scale, including samples from Mesoamerica. Inonotus linteus, an epitype of this species being designated here, is circumscribed as a narrow sense based on morphological and phylogenetic data, and this has lead to delimiting a new species I. cubensis Y.C. Dai, Decock&L.W. Zhou from Mesoamerica, and provides evidence for another two new additional species, I. alpinus Y.C. Dai&X.M. Tian and I. zonatus Y.C. Dai&X.M. Tian from East Asia. Phylogenetic analyses of ITS sequences also confirmed the status of I, baumii T. Wagner&M. Fisch.,I. linteus (Berk.&M.A. Curtis) Teixeira,I. lonicericola (Parmasto) Y.C. Dai,I. lonicerinus (Bondartsev) Sheng H. Wu, Y.C. Dai&T. Hatt.,I. vaninii (Ljub.) T. Wagner&M. Fisch.,I. sanghuang Sheng H. Wu, T. Hatt.&Y.C. Dai,I. weigelae T. Hatt.&Sheng H. Wu and I. weirianus (Bres.) T. Wagner&M. Fisch. as distinct taxa, while I. tenuicontextus L.W. Zhou&W.M. Qin is considered to be a synonym of I weigelae. A key to eleven species in the complex is provided.
For I. sanghuang growing exclusively on Morus, response surface methodology (RSM) was used to optimize the liquid fermentation medium. Plackett-Burman design (N=12) was used to examine the main factors affecting the mycelial biomass production from corn flour, glucose, peptone, yeast extract, bran, KH2PO4、MgSO47H2O and pH. The results showed that corn flour, yeast extract, bran and pH were selected as the main factors and the path of steepest ascent model was used to determine the optimum condition of mycelial biomass production. The four-factor, three-level Box-Behnken design (BBD) was employed to optimize the mycelial biomass. Under the optimized composition of culture medium (38.96g/L of corn flour,25.00g/L of glucose,3.75g/L of peptone,4.15g/L of yeast extract,20.55g/L of bran,1.25g/L of KH2PO4,0.625g/L of MgSO47H2O and pH6.39), the predicted and expected maximal mycelial biomass reached17.60g/L and18.33±0.86g/L, respectively. The optimized mycelial biomass was2.5times higher than the original mycelial biomass (7.29g/L).
The crude extracts of intercellular polysaccharide(IPS) and exopolysaccharides (EPS) produced by liquid fermentation of I. sanghuang were extracted by the way of water extraction followed by ethyl alcohol deposition. Two components (IPS1and IPS2) of IPS and two components (EPS1and EPS2) of EPS were isolated by a gel-filtration chromatography of Sephadex G-100. The results about the contents and DPPH radical scavenging activities of these six polysaccharides showed that there were no significant correlation between the polysaccharide contents and DPPH radical scavenging activities for all the six samples. The DPPH radical scavenging rates were enhanced with the rise of the polysaccharide contents of IPS, IPS1and IPS2in the concentration range of6.25μg/mL to500μg/mL. EPS, EPS1and EPS2showed no obvious DPPH radical scavenging activity in the test concentration range.
In order to probe into the effects of two-stage cultivation (consisted of an initial10days shake cultivation and then30days static culture) on the accumulation of I. sanghuang liquid fermentation metabolites, the contents change of total flavonoids, total phenolic compounds and polysaccharides in mycelia, as well as DPPH and hydroxyl radical scavenging activities of IS-EtOH and IS-H2O, were investigated during the whole period of two-stage cultivation. The results demonstrated that the contents of total flavonoids and total phenolic compounds in mycelia was found to increase dramatically by37.92%and77.27%, respectively, after10days and15days static culture. The results also implied that the richness of total flavonoids compounds increased dramaticly during the two-stage cultivation. The results of HPLC analysis were conjectured that rutin, eriodictyol, naringenin and sakuranetin may be contained in mycelia. The high free radical scavenging activity of IS-EtOH was resulted from the presence of phenolic-type compounds (total flavonoids and total phenolic compounds).
引文
白日霞,白日丽.碱提水溶针裂蹄多糖R的研究[J].天然产物研究与开发,1995,7(3)：41.
陈晓平,郝鑫,马吉霞.桑黄漆酶发酵条件的优化及其分离纯化[J].食品与发酵科技,2013,6：22-26.
程鑫颖,包海鹰,丁燕,等.瓦宁木层孔菌中5个化合物的抗肿瘤活性筛选[J].菌物研究,2011,9(3)：176-179.
褚以文.微生物培养基优化方法及其OPTI优化软件[J].国外医药(抗生素分册),1999,20(2)：58-60+66.
崔宝凯,戴玉成,杨宏.药用真菌粗毛纤孔菌概述[J].中国食用菌,2009,28(4)：6-7.
崔凤杰,王宏媛,杨焱,等.基于响应曲面法优化桑黄菌深层发酵条件[J].食品科技,2013,38(8)：61-66.
崔月花,彭祝明,胡嘉兴,等.8种渗水利湿中药对桑黄生长及多糖合成的影响[J].扬州大学学报(农业与生命科学版),2012,03：27-31.
戴玉成.药用担子菌—鲍氏层孔菌(桑黄)的新认识[J].中草药,2003a,34(1)：94-95.
戴玉成.一种新的药用真菌—瓦尼木层孔菌(杨黄)[J].中国食用菌,2003b,22(5)：7-8.
戴玉成.中国储木及建筑木材腐朽菌图志[M].北京,科学出版社,2009.
代志凯,张翠,阮征.试验设计和优化及其在发酵培养基优化中的应用[J].微生物学通报,2010,37(6)：894-903.
邓叔群.中国的真菌[M].科学出版社,北京,1963.
杜萍,陈艳秋.七个桑黄菌株酯酶同工酶的研究[J].中国食用菌,2006,25(2)：40-42.
杜萍,陈艳秋,李玉.鲍氏层孔菌基因组DNA提取及RAPD扩增条件优化[J].菌物学报,2007,26(2)：302-305.
樊锦艳,王秋颖,薛梅,等.桑黄胞外多糖生产培养基的初步研究[J].食品科技,2004,29(2)：93.
方白玉.桑黄菌丝液体培养条件初探[J].湖南师范大学自然科学学报,2008,31(1)：90-94.
高慧娟,刘雨晴,李娟辉,等.金属离子对桑黄菌丝体及胞外多糖含量的影响[J].食品研究与开发,2013,34(8)：20-23.
高凯,杜明,吕英华,等.10株桑黄菌基于rDNA ITS序列的分子鉴定[J].蚕业科学,2010,36(4)：0584-0589.
高兴喜,姚强,宫志远,郎晓,任鹏飞,刘岩.不同脂肪酸对桑黄液体发酵的影响[J].食品科学,2011,36(7)：198-201.
葛青,毛建卫,张安强,孙培龙.桑黄子实体多糖的分离纯化及结构鉴定[J].食品科技,2013,03：168-171+175.
郝瑞霞,周帅,杨焱,等.不同来源的桑黄子实体醇提物中黄酮含量及生物活性的比较[J].食用菌 学报,2008,15(2)：23-26.
何培新,刘伟.桑黄真菌的分子鉴定及分子系统学初探[J].湖北农业科学,2009,48(4)：778-780.
黄丽洋,石卉,王晓婷,等.野生桑黄菌株的分离、鉴定和次生代谢物分析[J].中草药,2013,44(23)：3394-3399.
黄年来,林志彬,陈国良,等.中国食药用菌学[M].上海：上海科学技术文献出版社.2010.
回晶,李辉,朱春玉,等.桑黄子实体与菌丝体营养成分的比较研究[J].特产研究,2009,(2)：59-61.
贾建波,李相前,杨文,张恒.桑黄多糖分离纯化及其结构初步鉴定[J].食品科学,2006,27(12)：446-450.
金城.食用菌多糖的抗氧化活性[J].微生物学通报,2013,40(4)：720.
兰进,徐锦堂,等.药用真菌栽培实用技术[M].中国农业出版社,2001.
雷萍,吴亚召,张飞龙,等.桑黄漆酶的发酵条件研究初报[J].中国食用菌,2012,31(5)：26-28.
雷萍,张文隽,吴亚召,等.桑黄菌液体发酵工艺研究[J].食用菌,2011,(3)：10-11.
李波,徐泽民,李方,等.试验设计与优化[J].中国皮革,2003,32(1)：26-28.
李宜明,沈业寿,季俊虬,等.桑黄菌质多糖体外抑瘤及抗环磷酰胺致突变的作用[J].中国科学技术大学学报,2006,36(7)：700-703.
林百全,余晓斌.利用SAS软件优化桑黄发酵培养基[J].食品与发酵工业,2006,32(11)：84-86.
刘惠知,邵晨霞,吴胜莲,等.桑黄菌的药理及产品开发研究进展[J].食用菌,2013,(5)：1-3.
刘艳芳,杨焱,贾薇,等.药用真菌桑黄总黄酮测定方法研究[J].食用菌学报,2006,13(2),49-52.
孟玲,王兰英,梁大勇,等.桦褐孔菌多糖测定方法的比较[J].食品研究与开发,2010,31(4)：108-110.
莫顺燕,杨永春,石建功.桑黄黄酮A和B的分离与合成[J].化学学报,2003a,61(7)：1161-1163.
莫顺燕,杨永春,石建功.桑黄化学成分研究[J].中国中药杂志,2003b,28(4)：54-56.
莫顺燕,杨永春,石建功.桑黄化学成分研究[J].中草药,2004,35(10)：21-23.
齐欣,张峻,陈颖,等.六种不同树种桑黄有效成分的比较[J].食品科学,2010,35(6)：199-201.
秦俊哲,李亚杰,帅斌.响应面法优化桑黄液体发酵培养条件的研究[J].食品科技,2010b,35(11)：7-10.
秦俊哲,王雅凤,帅斌.桑黄液体发酵培养基的响应面法优化研究[J].食品科技,2010a,35(3)：16-18+22.
秦俊哲,姚艳芳.桑黄菌的酯酶同工酶分析[J].食用菌,2006(6)：7-8.
邵杰,罗建光,曾晓雄.液体培养的桑黄胞外多糖发酵培养基成分的优化[J].食品科学,2012,37(3)：121-125.
沈业寿,郑立军,王正亮,等.桑黄胞内多糖抗肿瘤效应的实验研究[J].癌变畸变突变,2007,19 (4)：305-308.
史新强,沈业寿,卫自,等.桑黄胞内多糖对白血病细胞K562的增殖抑制效应研究[J].癌变畸变突变,2007,19(6)：440-441.
宋爱荣,王光远,赵晨,等.火针层孔菌(桑黄)粗多糖对荷瘤小鼠的免疫调节研究[J].菌物学报,2009,28(2)：295-298.
王丹,邹莉,杨民宝,等.药用真菌桑黄的系统发育分析[J].中国食用菌,2010,29(5)：40-42.
王清,沈业寿,赵浩如.桑黄子实体水提物抗肿瘤和抗环磷酞胺致突变作用研究[J].食用菌,2006,5：57-59.
王晓梅,罗友成,郭志勇,等.忍冬木层孔菌化学成分研究[J].中药材,2011,34(6)：891-893.
王中喜,王晓娟,田玉菊.桑黄多糖对小鼠骨髓造血功能损伤的保护作用[J].长春中医药大学学报,2006,22(2)：67.
魏江春.关于二十一世纪菌物系统生物学的目标与任务的思考[G].北京：中国菌物学会第三届会员代表大会暨全国第六届菌物学学术讨论会,2003,1-3.
魏磊,郑朝晖,侯成林,等.四种野生食用菌粗多糖的抗氧化活性[J].微生物学通报,2011,38(10)：1533-1539.
吴声华.珍贵药用菌“桑黄”物种正名[J].食药用菌,2013,20(3)：177-179.
吴声华,戴玉成,Hattori.桑黄真菌分类学研究[G].武汉：海峡两岸第十届菌物学暨第三届食药菌学术研讨会论文集,2011,91-92.
谢丽源,邓科君,张勇,等.桑黄深层发酵胞外酶活性的测定与分析[J].食品工业科技,2010a,31(7)：183-186.
谢丽源,谭伟,郭勇,等.响应面法优化桑黄产胞内多糖液体发酵培养基[J].食品科学,2011a,36(7)：224-228.
谢丽源,张勇,邓科君,等.基于rDNA ITS序列分析的桑黄真菌菌株分子鉴定[J].食品科学,2010c,31(9)：182-186.
谢丽源,张勇,郭勇,等.桑黄菌丝体多糖的分离纯化及理化性质分析[J].食品科学,2011b,36(5)：143-147.
谢丽源,张勇,彭金华,等.桑黄真菌分子鉴定及遗传多样性分析[J].菌物学报,2010b,29(3)：347-356.
杨全,张卉,王琦,等.桑黄胞外多糖抗肿瘤活性研究[J].北京中医药大学学报,2007,30(3)：188-190.
姚强,高兴喜,宫志远,等.部分稀土元素对桑黄液体发酵的影响[J].中国食用菌,2011,30(3)：41-44.
俞忠明,董宇,李洪玉,等.火木层孔菌桑黄醇提物的HPLC指纹图谱研究[J].中药材,2014,37(1)：41-44.
曾念开,王秋颖,苏明声.桑黄基原物种的探讨[J].中国食用菌,2008,27(2)：56-59.
张梅梅,魏志文,刘玉冰,等Folin-Ciocalteu比色法测定桦褐孔菌多酚的条件优化[J].菌物学报,2011,30(2)：295-304.
张楠,夏尚远,刘训理,等.统计优化技术在微生物发酵中的应用[J].山东农业大学学报,2009,40(3)：465-468.
张沙沙,陈莉,丁克坚,等.桑黄液体培养基的筛选及培养条件优化[J].东北林业大学学报,2013,41(6)：76-79.
张万国,胡晋红,蔡溱.桑黄对四氯化碳致大鼠肝损伤的保护作用[J].中国药房,2003,14(5)：267-269.
张小青.什么是桑黄?南通：中国菌物学会首届药用真菌产业发展暨学术研讨会论文集[G],2005,82.
张小青,戴玉成.中国真菌志(第二十九卷,锈革孔菌科)[M].北京：科学出版社,2005,101-165.
赵澜,张红峰.桑黄粗多糖对肿瘤细胞增殖及转移相关能力的抑制作用[J].华东师范大学学报,2008,3(2)：79-83.
赵子高,杨焱,刘艳芳,等.药用真菌桑黄液体深层发酵条件的优化[J].微生物学通报,2007,34(3)：459-463.
郑立军,沈业寿,季俊虬,等.桑黄胞内多糖的抗突变和抗氧化作用[J].癌变畸变突变,2006,18(6)：465-468.
郑立军,沈业寿,季俊虬,等.桑黄胞外多糖药理活性的初步研究[J].食品科学,2007,28(1)：318-321.
郑媛,沈业寿.桑黄胞内多糖抗衰老作用的研究[J].中国食用菌,2006,25(3)：38-41.
周海鸥,汪传高,张益波,等.统计学分析方法应用于桑黄菌发酵培养基的优化[J].食品研究与开发,2009,30(5)：44-48.
朱虎,何长川,张宗举,等.响应面法优化桑黄菌发酵培养基[J].中国生物制品学杂志,2011,24(6)：724-727.
邹祥,郭霞,孙敏.前体对桑黄菌丝生长和胞外多糖合成的影响[J].食品科学,2010,35(21)：262-265.
Amalfi M, Raymundo T, Valenzuela R, et al. Fomitiporia cupressicola sp. nov., a parasite on Cupressus arizonica, and additional unnamed clades in the southern USA and northern Mexico, determined by multilocus phylogenetic analyses[J]. Mycologia,2012,104(4):880-893.
Bai YH, Feng YQ, Mao DB, et al. Optimization for betulin production from mycelial culture of Inonot-us obliquus by orthogonal design and evaluation of its antioxidant activity[J]. Journal of the Taiwan Institute of Chemical Engineers,2012,43(5):663-669.
Berkeley MJ, Curtis MA. Characters of new fungi, collected in the North Pacific exploring expedition by Charles Wright[J]. Proceedings of the American Academy of Arts and Sciences.1860,4:111-13 0.
Box GP, Behnken DW. Some new three level design for the study of quantitative variables[J]. Technom-etrics,1960,2(4):456-475.
Bresadola G Selecta mycological 2, Stud trent,1926,7:51-58.
Brummitt PK, Powell CE. Authors of plant names[M]. Kew, Royal Botanic Gardens.1992.
Chen QH, He GQ, Mokhtar AMA. Optimization of medium composition for the production of elastase by Bacillus sp EL31410 with response surface methodology[J]. Enzyme and Microbial Technology, 2002,30(5):667-672.
Chen ZM, Li Q, Liu HM, et al. Greater enhancement of Bacillus subtilis spore yields in submerged cultures by optimization of medium composition through statistical experimental designs[J]. Appli-ed Microbiology and Biotechnology,2010.85(5):1353-1360.
Cho JY, Kwon YJ, Sohn MJ, et al. Phellinstatin, a new inhibitor of enoyl-ACP reductase produced by the medicinal fungus Phellinus linteus[J\. Bioorganic and Medicinal Chemistry Letters,2011,21 (6):1716-1718.
Chung KS, Kim SS, Kim HS, et al. An investigation on the antitumor constituents of Phellinus linteus[J]. Korean Journal of Mycology,1991,19:361.
Chung KS, Kim SS, Kim HS, et al. Effect of Kp an antitumor protein-polysaccharide from mycelial culture of Phellinus linteus on the humoral immune response of tumor bearing ICR mice to sheep red blood cells[J]. Archives of Pharmacal Research,1993,16(4):336-338.
Dai YC. Phellinus sensu lato (Aphyllophorales, Hymenochaetaceae) in East Asia[J]. Acta Botanica Fennica,1999,166:1-115.
Dai YC. Hymenochaetaceae (Basidiomycota) in China[J]. Fungal Diversity,2010a,45:131-343.
Dai YC, Cui BK, Tao WQ. Phellinus mori sp. nov. (Basidiomycota, Hymenochaetales) from northern China[J]. Mycotaxon,2008,105:53-58.
Dai YC, Xue MQ. Studies on the medicinal polypore, Phellinus baumii, and its kin, P. linteus[J]. Mycot-axon,1998,67:191-200.
Dai YC, Yang ZL, Cui BK, et al. Species diversity and utilization of medicinal mushrooms and fungi in China (Review)[J]. International Journal of Medicinal Mushrooms.2009,11:287-302.
Dai YC, Zhou LW, Cui BK, et al. Current advances in Phellinus sensu lato:medicinal species, functi-ons, metabolites and mechanisms[J]. Apply Microbiology Biotechnology.2010b,87(5): 1587-1593.
Dong W, Ning L, Lu WD, et al. Tumor-inhibitory and liver-protective effects of Phellinus igniarius extracellular polysaccharides[J]. World Journal Microbiology Biotechnology,2009,25(4):633-638.
Fang QH, Zhong JJ. Two-stage culture process for improved production of ganoderic acid by liquid fermentation of higher fungus Ganoderma lucidum[J]. Biotechnology Progress,2002,18(1):51-54.
Gilbertson RL, Ryvarden L. North American polypores 2. Megasporoporia-Wrightoporia[M]. Fungi-flora,1987, Oslo.
Guindon S, Gascuel O. A simple, fast and accurate method to estimate large phylogenies by maximum-likelihood[J]. Systerm Biology,2003,52(5):696-704.
Guo JJ, Zhu TB, Collins LA, et al. Modulation of lung cancer growth arrest and apoptosis by Phellinus Linteus[J]. Molecular Carcinogenesis,2007,46(2):144-154.
Hall TA. Bioedit:a user-friendly biological sequence alignment editor and analysis program for Windo-ws 95/98/NT[J]. Nucleic Acids Symp Ser (Oxf),1999,41:95-98.
Han KL, Lee Y, Song JH, et al. Enhanced production and secretion of rutin and GABA in immobilized cells of mulberry tree (Morus bombycis K.)[J]. Plant Cell Tissue and Organ Culture,2012, 108(3):513-520.
Han SB, Lee CW, Jeon YJ, et al. The inhibitory effect of polysaccharides isolated from Phellinus linteus on tumor growth and metastasis[J]. Immunopharmacology,1999,41(2):157-164.
Han SB, Lee CW, Kang JS, et al. Acidic polysaccharide from Phellinus linteus inhibits melanoma cell metastasis by blocking cell adhesion and invasion[J]. International Immunopharmacol,2006, 6(4):697-702.
He SH, Li HJ. Hymenochaete rhododendricola and H. quercicola spp. nov. (Basidiomycota, Hymenoc-haetales) from Tibet, southwestern China[J]. Nordic Journal of Botany,2011,29(4):484-487.
Humfeld H, Sugihara TF. Mushroom mycelium production by submerged propagation[J]. Food Techn-ology,1949,3:353-356.
Humfeld H, SugiharaTF. The nutrient requirements of Agaricus camestris grown in submerged cultu-re [J]. Mycologia,1952,44:605-621.
Hwang HJ, Kim SW, Choi JW, et al. Production and characterization of exopolysaccharides from submerged culture of Phellinus linteus[J]. Enzyme and Microbial Technology,2003(33):309-319.
Ikedawa T, Nakanishi M, Uehara N, et al. Anti-tumor action of some Basidiomycetes, especially Phellinus linteus[J]. Japanese journal of cancer research:Gann.,1968,59(2):155-157.
Jayaraman J, Jesudoss VAS, Menon VP, et al. Anti-inflammatory role of naringenin in rats with ethanol induced liver injury[J]. Toxicology Mechanisms and Methods,2012,22(7):568-576.
Jeon H, Hwang SG, Jung YH, et al. Inhibitory effect of oral administration of sangwhang mushroom (Phellinus linteus) grown on germinated brown rice on experimental lung metastasis and tumor growth in mice[J]. Food Science Biotechnol,2011,20(1):209-214.
Jeon TI, Hwang SG, Lim BO, et al. Extracts of Phellinus linteus grown on germinated brown rice suppress liver damage induced by carbon tetrachloride in rats[J]. Biotechnol Letter,2003,25(24): 2093-2096.
Jeong WJ, Lim YW, Lee JS, et al. Phylogeny of Phellinus and related genera inferred from combined data of ITS and mitochondrial SSU rDNA sequences[J]. Journal of Microbiol Biotechnol,2005, 15(5):1028-1038.
Keulen G, Jonkers HM, Claessen D, et al. Differentiation and anaerobiosis in standing liquid cultures of Streptomyces coelicolor[J]. Journal of Bacteriology,2003,185:1455-1458.
Kim GY, Han MG, Song YS, et al. Proteoglycan isolated from Phellinus linteus induces Toll-like recep tors 2-and 4-mediated maturation of murine dendritic cells via activation of ERK, p38, and NK2κB[J]. Biological & Pharmaceutical Bulletin,2004,27(10):1656-1662.
Kim GY, Lee JY, Lee JO, et al. Partial characterization and immunostimulatory effect of a novel polysa- ccharide-protein comp lexextracted from Phellinus linteus[J]. Biosci Biotechnol Biochem,2006, 70(5):1218-1226.
Kim GY, Park SK, Lee MK, et al. Proteoglycan isolated from Phellinus linteus activates murine B lymphocytes via protein kinase C and protein tyrosine kinase[J]. International Immunopharmacol, 2003,3(9):1281-1292.
Kim YS, Park KS, Park HK, et al. Compositional sugar analysis of antitumor polysaccharides by high performance liquid chromatography and gas chromatography[J]. Archives of Pharmacal Research, 1994,17(5):337-342.
Kong DH, Hong ND, Ha SB, et al. Stimulation of humoral and cell mediated immunity by po lysaccharide from mushroom Phellinus linteus[J]. International Journal of Immunopharmacol, 1996,18(5):295.
Kong KH, Kim HM, Hong DN, et al. The inhibitory effect of polysaccharide isolated from Phellinus linteus on tumor growth and metastasis[J]. Immunopharmacology,1999,41(2):157-164.
Kozarski M, Klaus A, Niksic M, et al. Antioxidative and immunomodulating activities of polysacchar-ide extracts of the medicinal mushrooms Agaricus bisporus, Agaricus brasiliensis Ganoderma lucidum and Phellinus linteus[J]. Food Chemistry,2011,129(4):1667-1675.
Lai MN, Ng LT, Antioxidant and antiedema properties of solid-state cultured honey mushroom, Armillaria mellea (higher Basidiomycetes), extracts and their polysaccharide and polyphenol contents[J]. International Journal of Medecinal Mushrooms,2013,15(1):1-8.
Larkin MA, Blackshields G, Brown NP, et al. Clustal W and clustal X version 2.0[J]. Bioinformatics,20 07,23(21):2947-2948.
Larsson KH, Parmasto E, Fisher M, et al. Hymenochaetales:a molecular phylogeny for the hymenoch-aetoid clade[J]. Mycologia,2006,98(6):926-936.
Lee JH, Cho SM. Effect of cultural conditions on polysaccharide production and its monosaccharide composition in Phellinus linteus,13202[J]. Korean Journal of Mycology,1995,23(4):325-331.
Lee JH, Cho SM, Yoo ID, et al. Immunostimulating activity an characterization of polysaccharides from Phellinus linteus[J]. Journal of Microbiol Biotechnol,1996,6(3):213-218.
Lee JW, Baek SJ, Bang KW, et al. Characteristics of polysaccharide isolated from the fruitbodies and cultured mycelia of Phellinus linteus IY001[J]. Korean Journal of Mycology,1999,27:424-429.
Lee Y, Lee DE, Lee HS, et al. Influence of auxins, cytokinins, and nitrogen on production of rutin from callus and adventitious roots of the white mulberry tree (Morus alba L.)[J]. Plant Cell Tissue Organ Culture,2011,105(1):9-19.
Lee YS, Kang IJ, Won MH, et al. Inhibition of protein tyrosine phosphatase lbeta by hispidin derivati-ves isolated from the fruiting body of Phellinus linteus[J]. Natural Product Communicatio-ns, 2010,5(12):1927-1930.
Li G, Kim DH, Kim TD, et al. Protein-bound polysaccharide from Phellinus linteus induces G2/M phase arrest and apoptosis in SW480 human colon cancer cells[J]. Cancer Letter,2004,216(2):175-181.
Lim WY, Lee JS, Jung HS. Type stydies on Phellinus baumii and Phellinus linteus[J]. Mycotaxon,2003, 85:201-210.
Liu CJ, Chiang CC, Chiang BH. The elicited two-stage submerged cultivation of Antrodia cinnamomea for enhancing triterpenoids production and antitumor activity[J]. Biochemical Engineering Journal, 2012,64:48-54.
Lyubarsky LV. Polypori nonnulli rari ex riente extremo[J]. Bot Mater Otd Spor Rast,1962,15:111-125.
Mao XB, Zhong JJ. Hyperproduction of cordycepin by two-stage dissolved oxygen control in submerg-ed cultivation of medicinal mushroom Cordyceps militaris in bioreactors[J]. Biotechnology Progress,2004,20(5):1408-1413.
Murrill WA. The Polyporaceae of North America 2. The genus Pyropolyporus[J]. Bull Torrey Club, 1903,30(2):109-120.
Nakamura T, Fukuda M, Matsugo S, et al. Analysis of mitochondrial DNA restriction fragment length polymorphisms for examining genetic variability among isolates of Phellinus linteus[J]. Mycoscie-nce,2002,43(6):443-445.
Naruse S, Takaeda S, Ito H, et al. Studies on antitumor activity of basidiomycetes:2. Antitumor effects of polysaccharides prepared from cultured basidiomycetes[J]. Mie Medical Journal,1974,23:207-230.
Oberwinkler F. Evolutionary trends in Basidiomycota[J]. Stapfia,2012,96:45-104.
Oh GT, Han SB, Kim HM, et al. Immunostimulating activity of Phellinus linteus extracts to B-lympho-cyte[J]. Archives of Pharmacal Research,1993,15(4):379-381.
Ostrakhovitch EA, Afanas'ev IB. Oxidative stress in rheumatoid arthritis leukocytes:suppression by rutin and other antioxidants and chelators[J]. Biochemical Pharmacology,2001,62(6):743-746.
Park HS, Kim GY, Nam BH, et al. The Determination of the Partial 28S Ribosomal DNA Sequences and Rapid Detection of Phellinus lintus and Related species[J]. Mycobiology,2002,30(2):82-87.
Park IH, Chung SK, Lee KB, et al. An antioxidant hispidin from the mycelial culture of Phellinus linteus[J]. Archives of Pharmacal Research,2004,27(6):615-618.
Parmasto E, Parmasto I. Phellinus baumii and related species of the Ph. linteus group (Hymenochaetac-eae, Hymenomycetes)[J]. Folia Cryptogamica Estonica,2001,38:53-61.
Patouillard PN. Quelques champignons de l'Afrique occidentale[J]. Bulletin de la Societe Mycology de France,1907,23:80-85.
Petersen JH. Farvekort. The danish mycological society's colour-chart [M]. Foreningen til Svampekun-dskabens Fremme,1996, Greve.6.
Plackett RL, Burman JP. The design of optimum multifactorial experiments[J]. Biometrika,1946,33(4): 305-325.
Posada D. jModelTest:Phylogenetic model averaging[J]. Molecular Biology and Evolution,2008, 25(7):1253-1256.
Rau U, Gura E, Olszewski E, et al. Enhance of glucan formation of filamentous fungi by effective mixing, oxygen limitation and fed-batch processing[J]. Journal of Indina Microbiology,1992, 9(1):19-26.
Rhee YK, Han MJ, Park SY. In vitro and in vivo anti-tumor activity of the fruitbody of Phellinus linteus[J]. Korean Journal of Food Science Technology,2000,32(2):477-480.
Ronquist F, Huelsenbeck JP. MrBayes 3:Bayesian phylogenetic inference under mixed models[J]. Bioi-nformatics,2003,19(12):1572-1574.
Sasaki T, Fujii K, Sugura M, et al. Antitumor polysaccharides from some polyporaceae, Ganderma applanatum (Pers.) Pat. and Phellinus linteus (Berk.& Curt.) Teng[J]. Chemical Pharmaceutical Bulletin,1971,19(4):821.
Sell I. Taxonomy of the species in the Phellinus igniarius group[J]. Mycotaxon,2008,104:337-347.
Shimizu T, Lin F, Hasegawa M, et al. The potential bioproduction of the pharmaceutical agent sakurane-tin, a flavonoid phytoalexin in rice[J]. Bioengineered,2012,3(6):352-357.
Shon YH, Nam KS. Antimutagenicity and induction of anticarcinogenic phase II enzymes by Basidio-mycetes[J]. Journal of Ethnopharmacology,2001,77(1):103-109.
Smith JE, Rowan NJ, Sullivan R. Medieinal mushrooms:a rapidly developing area of bioteclinology for cancer therapy and other bioactivities[J]. Biotechnology Letter,2002,24:1839-1845.
Song CH, Ra KS, Yang BK, et al. Immunostimulating activity of Phellinus linteus[J]. Korean Journal of Mycology,1998,26:86-90.
Song KS, Cho SM, KO KS, el al. Secondary melalmliles from the mycelial culture borth of Phellinus lnteus[J]. Agricultural, Chemistry and Biotechnology,1991,37(2):100.
Song KS, Li G, Kim JS, et al. Protein-bound polysaccharide from Phellinus linteus inhibits tumor growth, invasion, and angiogenesis and alters Wnt/β-catenin in SW480 human colon cancer cells[J]. BMC Cancer,2011,11:307.
Song YS, Kin SH, Sa JH, et al. Anti-angiogenic, antioxidant and xanthine oxidase inhibition activities of the mushroom Phellinus linteus[J]. Journal of Ethnopharmacology,2003,88(1):113-116.
Swofford DL. PAUP*:Phylogenetic analysis using parsimony (*and other methods). Version 4.0b10 [M]. Sinauer Associates,2002, Sunderland, Massachusetts.
TangYJ, Zhong JJ. Scale-up of a liquid static culture process for hyperproduction of ganoderic acid by the medicinal mushroom Ganoderma lucidum[J]. Bioteehnology Progress,2003a,19:1842-1846.
Tang YJ, Zhong JJ. Role of oxygen supply in submerged fermentation of Ganoderma lucidum for prod-uction of Ganoderma polysaceharide and ganoderic acid[J]. Enzyme Microbiology Technology, 2003b,32:478-484.
Tian XM, Yu HY, Zhou LW. et al. Phylogeny and taxonomy of the Inonotus linteus complex[J]. Fungal Diversity,2013,58(1):159-169.
Tomsovsky M, Vampola P, Sedlak P. et al. Delimitation of central and northern European species of the Phellinus igniarius group (Basidiomycota, Hymenochaetales) based on analysis of ITS and translation elongation factor 1 alpha DNA sequences[J]. Mycological Progress,2010,9(3):431-445.
Tseng YH, Yang JH., Mau JL. Antioxidant properties of polysaccharides from Ganoderma tsugae[J]. Food Chemistry,2008,107(2):732-738.
Tsuji T, Du W, Nishioka T, et al. Phellinus linteus extract sensitizes advanced prostate cancer cells to apoptosis in athymic nude mice[J]. Plosone,2010,5(3):9885.
Wagner T, Fischer M. Proceedings towards a natural classification of the world wide Phellinus and Inonotus s.l. and phylogenetic relationships allied genera[J]. Mycologia,2002,94(6):998-1016.
Wang Y, Yu JX, Zhang CL, et al. Influence of flavonoids from Phellinus igniarins on sturgeon caviar: antioxidant effects and sensory characteristics[J]. Food Chemistry,2012,131:206-210.
Wasser SP. Medicinal mushrooms as a source of antitumor and immunomodulating polysaccharides[J]. Apply Microbiology Biotechnology,2002,60:258-274.
Wasser SP. Current findings, future trends, and unsolved problems in studies of medicinal mushroom-ms[J]. Apply Microbiology Biotechnology,2011,89(5):1323-1332.
White TJ, Bruns TD, Lee S. et al. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In:Innis MA, Gelfand DH, Sninsky JJ, White TJ (eds) PCR protocols, a guide to methods and applications[M]. Academic, San Diego,1990.
Wu CY, Lin YP, Wang JC, et al. Effect of cultivation conditions on the producton of mycelial biomass and exopolysaccharide by submerged culture of a rooting shank medicinal mushroom, Oudemansiella radicata (Relhan) singer (Agaricomycetideae)[J]. International Journal of Medicin-al Mushroom,2010,12(1):99-110.
Wu SH, Dai YC, Hattori T, et al. Species clarification for the medicinally precious'sanghuang'mushr-oom[J]. Botanical Studies,2012,53(1):135-149.
Xiang YL, Xu XQ, Li J. Chemical properties and antioxidant activity of exopolysaccharides fractions f-rom mycelial culture of Inonotus obliquus in a ground corn stover medium[J]. Food Chemistry, 2012,134(4):1899-1905.
Xue Q, Sun J, Zhao MW, et al. Immunostimulatory and anti-tumor activity of a water-soluble polysac-charide from Phellinus baumii mycelia[J]. World Journal of Microbiology Biotechnology,2011, 27(5):1017-1023.
Yayeh T, Oh WJ, Park SC, et al. Phellinus baumii ethyl acetate extract inhibits lipopolysaccharide-ind-ced iNOS, COX-2, and proinflammatory cytokine expression in RAW264.7 cells[J]. Journal of Natural Medicine,2012,66(1):46-54.
Yeou LM, Chung TJ, Chang HP. Antioxidant properties of edible basidiomycete Phellinus igniarius in submerged cultures[J]. Journal of Food Science,2010,75(1):18-24.
Zhang WY, Lee JJ, Kim Y, et al. Effect of eriodictyol on glucose uptake and insulin resistance in vitro[J]. Journal of Agricultural and Food Chemistry,2012,60(31):7652-7658.
Zhao CL, Cui BK. A new species of Perenniporia (Polyporales, Basidiomycota) described from southe-rn China based on morphological and molecular characters [J]. Mycological Progress,2012, 11(2):555-560.
Zhao L, Fan F, Wang P, et al. Culture medium optimization of a new bacterial extracellular polysaccha-ride with excellent moisture retention activity[J]. Apply microbiology Biotechnology,2013, 97(7):2841-2850.
Zhou LW, Dai YC. Phylogeny and taxonomy of Phylloporia (Hymenochaetales) with the description of five new species and a key to worldwide species[J]. Mycologia,2012,104(1):211-222.
Zhou LW, Jia BS. A new species of Phellinus (Hymenochaetaceae) growing on bamboo in tropical China[J]. Mycotaxon,2010,114:211-216.
Zhou LW, Qin WM. Inonotus tenuicontextus sp. nov. (Hymenochaetaceae)from Guizhou, southwest China with a preliminary discussion on the phylogeny of its kin[J]. Mycological Progress,2011, 11(3):791-798.
Zhou LW, Xue HJ. Fomitiporia pentaphylacis and F. tenuitubus spp. Nov. (Hymenochaetales, Basidio-mycota) from Guangxi, southern China[J]. Mycological Progress,2012, 11(4):907-903.
Zhu H, Liu W, Wang SX, et al. Evaluation of anti-quorum-sensing activity of fermentation metabolites from different strains of a medicinal mushroom, Phellinus igniarius[J]. Chemotherapy,2012,58 (3):195-199.