Abstract
Background: Endophytic fungi are largely underexplored in the discovery of natural bioactive products though being rich sources of novel compounds with promising pharmaceutical potential. In this study, Taxus wallichiana, which has huge medicinal value, was investigated for its endophytic diversity and capability to produce bioactive secondary metabolites by analyzing antioxidant, antimicrobial and cytotoxic properties. Methods: The endophytes were identified by ITS-PCR using genomic DNA samples. The secondary metabolites were extracted by solvent extraction method using ethyl acetate. The antioxidant activity was analyzed by Thin Layer Chromatography, Total Phenol Content (TPC), Total Flavonoid Content (TFC) and DPPH assay, and the antimicrobial activity was analyzed by agar-well diffusion method. Brine shrimp lethality assay was used to analyze the cytotoxicity of the fungal extracts. Results: Out of 16 different Taxus trees sampled from different locations of Dhorpatan, 13 distinctive endophytic fungi were isolated and grouped into 9 different genera: Bjerkandera, Trichoderma, Preussia, Botrytis, Arthrinium, Alternaria, Cladosporium, Sporormiella and Daldinia. The ethyl acetate extracts isolated from three endophytic fungi: Alternaria alternata, Cladosporium cladosporioides and Alternaria brassicae showed significant TPC values of 204±6.144, 312.3±2.147 and 152.7±4.958µg GAE/mg of dry extract, respectively, and TFC values of 177.9±2.911, 644.1±4.202 and 96.38±3.851µg RE/mg of dry extract, respectively. Furthermore, these three extracts showed a dose dependent radical scavenging activity with IC 50 concentration of 22.85, 22.15 and 23.001 µg/ml, respectively. The extracts of C. cladosporioides and A. brassicae also showed promising antimicrobial activity against Escherichia coli, Staphylococcus aureus and Bacillus subtilis with a minimum inhibitory concentration of 250μg/ml for all bacteria. Both the samples showed cytotoxic property against shrimp nauplii with LC 50 of 104.2 and 125.9µg/ml, respectively. Conclusions: The crude fungal extracts obtained from endophytes: A. alternata, C. cladosporioides and A. brassicae upon purification and further identification of the bioactive compounds can be a fascinating source for novel pharmaceutical agents.
Highlights
Taxus wallichiana Zucc. (Himalayan yew) is an evergreen coniferous tree (10 to 28 meters in height), native to the Himalaya and parts of southeast Asia that grows on steep, moist mountain slopes at altitudes of about 2000m to 3500m above sea level
Only a few plants are investigated for their endophytic biodiversity and their potential to produce bioactive secondary metabolites
Nepal has an abundance of medicinal plants, so this research helps shed a light in the possibilities of discovering novel compounds with pharmaceutical potential from a variety of endophytic fungi residing within them
Summary
Taxus wallichiana Zucc. (Himalayan yew) is an evergreen coniferous tree (10 to 28 meters in height), native to the Himalaya and parts of southeast Asia that grows on steep, moist mountain slopes at altitudes of about 2000m to 3500m above sea level. Taxus wallichiana, which has huge medicinal value, was investigated for its endophytic diversity and capability to produce bioactive secondary metabolites by analyzing antioxidant, antimicrobial and cytotoxic properties. The ethyl acetate extracts isolated from three endophytic fungi: Alternaria alternata, Cladosporium cladosporioides and Alternaria brassicae showed significant TPC values of 204±6.144, 312.3±2.147 and 152.7±4.958μg GAE/mg of dry extract, respectively, and TFC values of 177.9±2.911, 644.1±4.202 and 96.38±3.851μg RE/mg of dry extract, respectively. These three extracts showed a dose version 2 (revision)
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