摘要
植物内生菌作为一种新的微生物资源逐渐成为人们寻找新天然产物的热点。植物内生真菌作为植物内生菌的重要成员，由于它的次级代谢产物往往种类繁多、骨架新颖、生物活性多样，已成为发现新的生物活性物质的重要来源。
本论文以三种不同药用植物(滑桃树、喜树和番荔枝)为研究对象，对来源于这三种药用植物的四株内生真菌(两株番荔枝植物内生真菌Pyrenochaeta sp．B36、Xylaria sp．A45，一株滑桃树植物内生真菌Aspergillus sp．F1和一株喜树内生真菌Phomopsis sp．XZ26)的次级代谢产物进行分离纯化和结构鉴定。共分离并鉴定了52个化合物(包括2个乙酰化产物和1个乙酯化产物)，其中23个为新化合物(反应产物不属于新化合物)，对其中部分化合物进行抗菌、抗肿瘤活性研究。由于目前对这三种药用植物内生真菌的次级代谢产物的研究较少，许多化合物都是首次从它们的内生真菌中分离得到。
从菌株Aspergillus sp．F1的次级代谢产物中分离并鉴定了12个化合物，包括聚酮类化合物、萜类化合物、部分萜类化合物。其中LT24为新化合物，命名为butyl 3，5-dichloro-2-hydroxy-6-(2-hydroxy-4-methoxy-6-(methoxycarbonyl)phenoxy)-4-methylbenzoate；新化合物LT 1为部分萜类化合物，命名为butyrolactone-V。
运用核磁共振、质谱及X-Ray单晶衍射分析技术，从菌株Phomopsis sp．XZ26的液体发酵和固体发酵产物中分离并鉴定了14个化合物，包括11个聚酮(LTC，LTP，LTQ，LTJ-1，LTJ-2，LTG，LTH，26-L-2，LTL，LTM，LTR)类化合物、2个(LTA，LTO)萜类化合物和1个(LTD)大环内酯类化合物。其中化合物LTC、LTG、LTH、LTJ-1、26-L-2、LTL、LTO、LTM、LTQ、LTR为新化合物；化合物26-L-2，LTG，LTL，LTM和LTR是首次从喜树内生真菌的次级代谢产物中分离得到，对其中部分化合物进行了抗菌活性测定。结果表明，这些化合物具有一定的抗菌活性。
从菌株Pyrenochaeta sp．B36的次级代谢产物中分离并鉴定了10个(B36-1，LT-36-2，LT-36-4，LT-36-9，LT-36-11，LT-36-12，LT-36-15，LT-36-16，LT-36-17，LT-36-19)聚酮类化合物。其中化合物LT-36-2，LT-36-12，LT-36-15，LT-36-16和LT-36-11是新化合物，且首次从番荔枝内生真菌中分离得到。化合物B36-1是聚酮类化合物，有很强的抗肿瘤活性：在浓度为5μg／ml时，对Raji细胞的抑制率为82．21％；在浓度为1．25μg／ml时，对HEPG2的抑制率为49．79％。
在对番荔枝另外一株内生真菌Xylaria sp．A45的研究中，从其PDA平板发酵产物中共分离并鉴定了13个化合物，包括9个聚酮类化合物(LT-45-1，LT-45-2，LT-45-4，LT-45-5，LT-45-7，LT-45-8，LT-45-10，LT-45-12，LT-45-14)、一个生物碱类化合物(LT-45-24)和三个萜类化合物(LT-45-16，LT-45-21和LT-45-25)，其中LT-45-2、LT-45-10、LT-45-16、LT-45-21、LT-45-25、LT-45-24为新化合物。
本论文的结果表明，植物内生真菌作为植物生态一个重要的组成部分，其次级代谢产物具有结构新颖性与生物活性多样性的特征，是寻找药物先导化合物的重要资源。
Recently,endophyte as a new microbiology resource has played an importantrole in people discovering novel natural products.Endophytic fungi is an importantpart of the endophyte,they can produce numerous secondary metabolites with severalstructures,novel skeletons and varied and high bioactivities.They have been a veryimportant source for searching new bioactivity natural product over decades.
In this thesis,three medical plants (Trewia nudiflora,Camptotheca acuminata,Annona squamosa L.) were picked as research sources.The secondary metabolites offour endophytic strains of different plants (two strains Pyrenochaeta sp.B36 andXylaria sp.A45 from Annona squamosa L.,one strain Aspergillus sp.F1 from Trewianudiflora and one strain Phomopsis sp.XZ26 from Camptotheca acuminata ) wereinvestigated.Fifty two compounds were isolated and elucidated from these fourstrains,including twenty three new compounds.Partial compounds were tested for theantimicrobe and anticancer bioactivity.There were not many research about thesemedical plants's endophytic fugi,so many compounds were isolated for the first time.
Twelve compounds were isolated from the strain Aspergillus sp.F1.Theirstructures were elucidated by extensive spectroscopic data analyses,includingpolyketides,terpenoids and dibenzylbutyrolactone lignans.One of them was new polyketide,namely LT-24,named butyl 3,5-dichloro-2-hydroxy-6-(2-hydroxy-4-methoxy-6-(methoxycarbonyl)phenoxy)-4-methylbenzoate,another one was newdibenzylbutyrolactone lignan,namely LT1,named butyrolactone-V.
The secondary metabolites of the strain Phomopsis sp.XZ26 for both solid andliquid fermentations were studied.Fourteen compounds were isolated.Theirstructures were elucidated by extensive spectroscopic data analyses and X-raycrystallographic analysis,including eleven (LTC,LTP,LTQ,LTJ-1,LTJ-2,LTG,LTH,26-L-2,LTL,LTM,LTR) polyketides,two (LTA,LTO) terpenoids and one (LTD)macrocyclic lactone.Among them,ten new compounds (LTC、LTD、LTG、LTH、LTJ-1、26-L-2、LTL、LTO、LTM、LTQ、LTR)were obtained.Except LTO was monoterpen,the others were all new polyketides.Compounds 26-L-2,LTG,LTL,LTM and LTR were polyketides isolated from Camptotheca acuminata fungi for thefirst time.Some of the compounds were tested for anti-microbial activities.The resultshowed that they had humble activities against Escherichia coli,Bacillus subtilis,Staphylococus aureus,Sacharomyces cereviiae and Candidia albicans.
Ten polyketides were isolated and identified from the strain Pyrenochaeta sp.B36 (B36-1,LT-36-2,LT-36-4,LT-36-9,LT-36-11,LT-36-12,LT-36-15,LT-36-16,LT-36-17,LT-36-19).LT-36-2,LT-36-12,LT-36-15,LT-36-16 and LT-36-11 were newcompounds and they were isolated from Annona squamosa L.fungi for the first time.Compound B36-1 was polyketide.When tested with anticancer bioactivity,thiscompound exhibited very strong activity.At the concentration of 5μg/ml,it still has82.21% inhibition rate against the Raji cell.When tested against HEPG2 cell,at theconcentration 1.25μg/ml,it still showed inhibition rate at 49.79%.
From another Annona squamosa L.'s endophytic fugus Xylaria sp.A45,thirteencompounds were obtained,including nine polyketides (LT-45-1,LT-45-2,LT-45-4,LT-45-5,LT-45-7,LT-45-8,LT-45-10,LT-45-12,LT-45-14),three new steroids(LT-45-16,LT-45-21 and LT-45-25) and one alkoid (LT-45-24).Compounds LT-45-2,LT-45-10,LT-45-16,LT-45-21 and LT-45-25 were new compounds.
In conclusion,this thesis indicated that endophytic fungi,this important part ofplant system,contain novel and bioactive compounds in its secondary metabolisms.Itis an important source for searching lead compounds for drugs.
引文
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