| Plant endophytic fungi, which included many different species and distributed widely, have been recognized as the richest sources for the discovery of secondary metabolites with a diverse range of unique structure and biological activity. The traditional research strategies for screening fungal secondary metabolites such as active-guide, chemical-based or their combination, often resulting in rediscovery of the previously described strains and already known compounds. With the development of molecular biology, polyketide synthase functional genes (PKSs)-based analysis has become to be one of the most important strategies to screen novel compounds.Salicornia bigelovii Torr., which thrives in the high-salt environment, can produce diverse bioactive compounds with unique skeletons. And studies have also shown that S. bigelovii Torr. hosts many endophytes which have the capability to produce bioactive secondary metabolites.In this study, fresh S. bigelovii Torr. was extracted by using ethyl acetate, and four pure compounds were obtained from the ethyl acetate extract of S. bigelovii Torr. by variety of separation and purification methods, such as silica gel column chromatography and Sephadex LH-20. Then the four compounds were determined to be oleic aicd (1)、 (3β,5a,8a,22E)-5,8-epidioxyergosta-6,9,22-trien-3-ol (2)、pentadecyl ferulate (3) and scopoletin (4) by modern spectroscopic techniques as well as comparison with related literature search.Two endophytic fungi named Salicorn 54 and Salicorn 46 were isolated and purified from S. bigelovii Torr. by the PDA plate and the slant culture and then assessed by PCR for the presence of PKS I genes. Result showed that a PKS gene of strain Salicorn 46 was screened. Based on BLAST analyses, the PKS I sequence from strain Salicorn 46 shared high sequence similarity (90-95%) with other previously reported sequences related to biosynthetic pathways. The phylogenetic tree showed that the unique KS sequence from the strain Salicorn 46 is clearly separated from any known sequence and could be involved in as yet uncharacterized pathways, thus indicating this isolate might have the potential for producing novel polyketides.The two endophytic fungi, Salicorn 54 and Salicorn 46, were identified as Phoma betae and Penicillium citrinum by using molecular biology analysis and morphological observation, respectively.The strain Salicorn 46 was selected for further cultivation, and eight pure compounds were isolated by the solvent extraction and a range of separation and purification methods, and the compounds were determined to be penicitriketo (5), ergone (6), (3p,5a,8a,22E)-5,8-epidioxyergosta-6,9,22-trien-3-ol (7), (3p,5a,8a,22E)-5,8-epidioxyergosta-6,22-dien-3-ol (8), stigmasta-7,22-diene-3β,5α,6α-triol (9),3β,5a-dihydroxy-(22E,24R)-ergosta-7,22-dien-6p-yl oleate (10), Nb-acetyltryptamine (11) and 2-(1-oxo-2-hydroxy-ethyl) furan (12) by nuclear magnetic resonance, infrared, ultraviolet, mass spectrometry, gas chromatography-mass spectrometry and other modern spectroscopic techniques.DPPH radical scavenging method was used to primarily evaluate the antioxidant activities of the 12 compounds. Result showed that all the tested compounds had the DPPH radical-scavenging activities to some extent and compounds 3 and 5 possessed the strongest DPPH radical scavenging activities, which were almost similar to VE (positive control).The obtained compounds were tested for their antimicrobial activity against eight pathogenic strains. Result revealed that compunds 6-10 had antimicrobial activities with different levels. And compound 6 had broad-spectrum antimicrobial activities against Candida albicans, Clostridium perfringens, Mycobacterium smegmatis, and M. phlei with MIC values 25.5 ± 0.48,25.5±0.25,18.5±0.34, and 51.0±0.68μM, respectively. Compound 7 displayed potent antimicrobial activities against C. perfringens and M. tetragenus with the same MIC values 23.5μM. Compounds 9 and 10 showed high levels of selectivity towards B. subtilis and M. phlei with MIC values 22.5±0.45 and 14.4±0.34 μM, respectively. |
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