Metabolites of endophytic fungi from Australian native plants as potential anticancer agents.

Interest in endophytes as natural sources for new medicines was inspired by the discovery of paclitaxel-producing endophytic fungi. This study investigated the anti-cancer activity of extracts of endophytes isolated from two Australian plants, Eremophila longifolia (EL) and Eremophila maculata (EM). Endophytes were isolated from surface-sterilised leaf tissue, grown as pure cultures and identified by sequencing of Internal Transcribed Spacer (ITS) regions of the ribosomal DNA. To determine cytotoxicity, two leukaemic (MOLT-4, T-cell leukaemia; PreB-697, Pre-B leukaemia), a lung cancer cell line (A549) and a normal human fibroblast cell line were treated with endophyte extracts to assess cytotoxicity in relation to alternariol monomethyl ether (AME) and alternariol (AOH). Endophyte extracts that showed cell cytotoxicity were analysed by UV-HPLC to determine the metabolites. Pure AME and AOH, three extracts form Alternaria sp. (EM-6, EM-7 and EM-9) and one from Preussia minima (EL-14) were cytotoxic to the cancer cell lines. All cytotoxic endophytes contained AME and AOH, the most cytotoxic endophyte EM-6 also contained two unique peaks. These data indicate that these four endophyte extracts may have anti-cancer properties due to the presence of AME and AOH; however, the unique compounds found in the EM-6 extract may be exclusively cytotoxic and warrant further investigation.

Optimization of protease production by endophytic fungus, Alternaria alternata, isolated from an Australian native plant.

Endophytes are recognised as potential sources of novel secondary metabolites, including enzymes and drugs, with applications in medicine, agriculture and industry. There is a growing need for new enzymes, including proteases, for use in industry that can function under a variety of conditions. In this study, three fungal endophytes (Alternaria alternata, Phoma herbarum and an unclassified fungus), were isolated from the Australian native plant, Eremophilia longifolia, and assessed for production of proteases. The lyophilised growth media obtained after fungal fermentation were analysed for protease production using enzyme activity assays. Protease production was optimised by assessing the effects of temperature, pH, carbon source and nitrogen source on activity. A. alternata showed the greatest protease activity in a wide range of pH (3-9). The broadest activity between 9 and 50 °C was observed at pH 7, suggesting a neutral protease. Overall, the optimum conditions were 37 °C and pH 7 with a maximum specific activity value of 69.86 BAEE units/mg. The characteristics demonstrated by this fungal endophyte showed that it is a potential source of an enzyme with particular application in the dairy industry. However, further studies of the tolerance to higher temperatures and pH will indicate whether the enzyme is suitable to such applications.

Endophytes from an Australian native plant are a promising source of industrially useful enzymes.

Endophytes are microorganisms that live within plant tissues that are potential sources of novel bioactive compounds, including enzymes. We have identified endophytes of the Australian native plant Eremophilia longifolia which were screened for the production of industrially useful enzymes. Seventeen fungal endophytes were isolated from the leaves of E. longifolia and enzyme production was investigated within a range of pH (3.5, 5.5, 7 and 9) and temperatures (9, 25, 37 and 50 °C). Amylase was the most common enzyme encountered with numerous isolates showing production throughout the temperature and pH ranges. Protease production was also seen over the conditions tested but was more dominant at lower pH and temperature. Activity was not observed for other enzymes including ligninase, xylanase and cellobiohydrolase. Enzymes from isolates of Preussia minima, Alternaria sp. and an unclassified fungus, which showed highest activity in screening assays, were investigated further. Enzyme production was verified by zymography and the amylase activity of P. minima was found to be significantly greater than that of Aspergillus oryzae particularly in alkaline conditions and low temperature which are desirable properties for the detergent industry. This work shows that enzymes with potential use in industry can be readily identified in fungal endophytes.

Four eremophilane sesquiterpenes from the mangrove endophytic fungus Xylaria sp. BL321.

Three new eremophilane sesquiterpenes (1-3) were isolated from the mangrove endophytic fungus Xylaria sp. BL321 together with 07H239-A (4), a known analogue of the new compounds. The structures of these compounds were elucidated by analysis of their MS, 1D and 2D NMR spectroscopic data. Compound 4 showed activation activity on α-glucosidase at 0.15 µM (146%), and then, 4 gradually produced inhibitory activity on α-glucosidase with increasing concentration, and the IC50 value is 6.54 µM.