Secondary Metabolites From Chaetomium Globosum CDW7, An Endophytic Fungus In Ginkgo Biloba

Endophytic fungi have been recognized as a potential source of bioactive secondary metabolites. Ginkgo biloba, one of the oldest species on earth with fossil records dating back more than200million years, has long been used in China as a traditional medicine for various ailments. Previous studies reported that Ginkgo biloba had fewer pests and diseases than other gymnosperms, which may be caused in part by its associated endophytes because many endophytes were able to synthesize bioactive compounds to defend host plants against pathogenic fungi and bacteria.Here we performed anti-fungal bioassay of eighty endophytic fungi isolated from Ginkgo biloba. Fifteen endophytes were active against at least one of the selected fungi, Fusarium graminearum, Sclerotinia sclerotiorum and Phytophthora capsici, by using agar diffusion method. The most bioactive strain CDW7was identified as Chaetomium globosum by microscopic examination and ITS rDNA sequence data. Three-fold-diluted culture broth of CDW7completely inhibited the mycelial growth and conidia germination of Fusarium graminearum in vitro. Therefore, Fusarium head blight (FHB), a common disease in wheat and barley associated with Fusarium spp., was used to test the anti-phytopathogenic activity in vivo. The fermentation broth of CDW7resulted in a protective efficacy of54.9%and curative efficacy of48.8%. Followed by a bioassay-guided approach, six compounds were obtained including two new chaetoglobosins21-methoxy-chaetoglobosin A (LB-1) and chaetoglobosin Y (LB-4) together with four known compounds chaetoglobosins A (LB-3), C (LB-5), F (LB-2), and1,2-benzenedicarboxaldehyde-3,4,5-trihydroxy-6-methyl (flavipin, LB-6). Flavipin was demonstrated to significantly inhibit the growth of several plant-pathogenic fungi, especially Fusarium graminearum with an EC50value of0.73μg/mL comparable to the commonly used fungicide carbendazim, indicating that it could be used as a fungicide or a lead compound of new fungicides. Chaetoglobosins A also showed consideribly antifungal activity with an EC50value below1.0μg/mL against Sclerotinia sclerotiorum, Fusarium graminearum, Phytophthora capsici and Alternaria solani.In our study, endophytic fungi from Ginkgo biloba were also screened for their antioxidant abilities by methods of DPPH radical-scavenging capacity assay (DPPH) and ferric-reducing antioxidant power assay (FRAP). Seven endophytic fungi were able to synthesize antioxidant substances and identified by ITS rDNA sequences. Among them, Chaetomium globosum CDW7displayed significant antioxidant capacity and flavipin was yielded as the most bioactive metabolite. Both in vitro (DPPH, ABTS, FRAP, ORAC, LPO) and in vivo (SOD, GSH-Px, MDA) tests showed that flavipin possessed the potential to be developed as a new antioxidant.Since strain CDW7was a remarkable producer of flavipin, the fermentation parameters of CDW7was then optimized for high flavipin production. Cultured under the optimal condition (25℃,100/250mL flask,12discs/flask,150rpm, pH6.5) for14days in PD broth medium, CDW7was able to synthesize flavipin at a production of315.5mg/L. In addition, flavipin output was positively correlated to antioxidant activities of crude extracts with a determination coefficient of0.8235, indicating that flavipin was the major antioxidant component of CDW7’s metabolites. These data demonstrated that flavipin is a potent antioxidant which has great potential in the therapy for free radical-associated diseases and CDW7was a highly-yielded bio-source of antioxidant flavipin.