The Secondary Metabolites Of Endophytic Fungi J14 From Nerium Indicum

Endophytic fungi are the fungi that colonize and cause asymptomatic infections in healthy plant tissues. There is a certain mutual symbiotic relationship between endophytic fungi and plants. Endophytic fungi can produce a variety of secondary metabolites which are identical or similar to plant components and have diverse biological activities and a great value in medicine and agriculture industries such as promoting plant growth, antimicrobial activities, antioxidant activities, antitumor activities, insecticidal activities and so on. Therefore, that is significant to study of the active ingredients of endophytic fungi.The endophytic fungus J14 which was isolated from Nerium indicum with higher antimicrobial activities was studied. According to its morphological characteristics and ITS sequence, the strain J14 was defined as the genus Alternaria (the homology was 99% between the strain J14 and Alternaria sp. SPS-04). Solid state fermentation was used to ferment the strain J14 in rice medium and the fermentation products were extracted with petroleum ether, ethyl acetate and methanol, concentrated to get 460 g of metabolite extracts.The metabolite extract was segregated by the silica gel column, sephadex LH-20 column chromatography, recrystallization, and 15 compounds were obtained. Their structures were determined by their physicochemical properties, ESI-HRMS, 1H-NMR and 13C-NMR data. These compounds were determined as: alternariol methyl ether (1), ergosterol peroxide (2), alternariol (3), malformin A1 (4), thymine (5), uracil (6), erythritol (7), xanthine (8), hypoxanthine (9), adenine (10),1,6-di-O-acetyl-mannitol (11), allantoin (12), thymidine (13), uridine (14), mannitol (15). Among them, compounds 4,5,6,8,9,11,12,13,14 were firstly obtained from fermentation products of the genus Alternaria. This paper reported 13C-NMR data of 1,6-di-O-acetyl-mannitol (11) firstly and NMR data of compound 11 were assigned.Antimicrobial activities of 15 compounds were tested against 4 bacteria (Staphylococcus aureus, Streptococcus lactis, Enterococcus coli, Pseudomonas aeruginosa) and 10 plant pathogenic fungi(Fusarium graminearum, Botrytis cinerea, Alternaria alternata, Alternaria brassicae, Phytop hthoracapsic, Valsa mali, Colletotrichum gloeosporioides, Peony anthracnose, Setosphaeria turicia, Sclerotinia sclerotiorum), and sodium penicillin, streptomycin sulfate, carbendazim were selected as the positive reference samples. The results showed that compound 1 (alternariol methyl ether) and compound 3 (alternariol) showed excellent antimicrobial activities as same as the positive reference samples. Compound 4 (malformin A1) and compound 12 (allantoin) showed mild antimicrobial activities.Antioxidant activities of 15 compounds were tested, and vitamin C was selected as the positive reference sample. The results showed that compound 6 (uracil), compound 10 (adenine) and compound 12 (allantoin) showed a certain degree of antioxidant activity.The study of the secondary metobalities of the endophytic fungi J14 which was isolated from Nerium indicum showed that the strain J14 could produce the coumarins which had broad antimicrobial activities and the alkaloids which had a certain degree of antioxidant activity. Therefore, the strain J14 would become a new resource for antimicrobial and antioxidant drugs.