Discovery of the anticancer drug vinblastine from the endophytic Alternaria alternata and yield improvement by gamma irradiation mutagenesis.

AIMS: Several fungal endophytes were isolated from some medicinal plants and screened for their ability to produce the anticancer drug vinblastine. METHODS AND RESULTS: An isolate was found to produce vinblastine (205·38 µg l-1 ), and the identity of the fungal vinblastine was confirmed by UV spectroscopic, high-performance liquid chromatography and electrospray ionization mass spectrometry analyses. Based on both morphological and molecular studies, the vinblastine-producing strain was identified as Alternaria alternata. Cytotoxic activities of the fungal vinblastine were evaluated against CHO-K1, MCF-7 and HepG-2 cell lines by the MTT assay. The proliferation of these cell lines was inhibited after treatment with fungal vinblastine and the recorded IC50 values of the respective cell lines were 12·15, 8·55 and 7·48 µg ml-1 . A strain improvement programme for improving vinblastine productivity by the fungal strain was also used. In addition, 10 broth media were evaluated for further increasing the production of vinblastine. The yield of vinblastine was intensified by 3·98-fold following gamma irradiation at 1000 Gy, and a stable mutant strain was isolated. Among the screened media, M1D broth (pH 6·0) stimulated the highest vinblastine production of 1553·62 µg l-1 by the isolated mutant strain. CONCLUSIONS: The present study is the first report on the production and yield improvement of the anticancer drug vinblastine by A. alternata. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings suggest A. alternata as a viable and potent source with excellent biotechnological potential for the production of vinblastine.

Agro-industrial wastes for production of paclitaxel by irradiated Aspergillus fumigatus under solid-state fermentation.

AIMS: Paclitaxel is the most profitable drug ever developed in cancer chemotherapy; however, the yield of paclitaxel from microbial platforms is still far from the commercial purpose. Thus, this study was conducted to explore the possibility of solid-state fermentation (SSF) for production of paclitaxel by fungal fermentation. METHODS AND RESULTS: Different agro-industrial wastes were screened as solid substrates for production of paclitaxel by the endophytic Aspergillus fumigatus TXD105 under SSF. Sugarcane bagasse followed by wheat bran, and rice bran were the most suitable substrates for maximum production of paclitaxel. In the effort to increase the paclitaxel production, selection of the most proper moistening agent that supports the production of paclitaxel by the fungal strain was investigated. The effect of varying inoculum concentrations on the production of paclitaxel was also studied. Moreover, optimization of SSF conditions (moisture level, substrate concentrations and nutrients concentration) was adopted using response surface methodology. SSF carried out under the optimum conditions of 20 g sugarcane bagasse, twofold nutrients concentration of the MM1D broth, 80% moisture level and inoculum concentration of 107 spores per ml intensified the paclitaxel concentration to 145·61 mg kg-1 which represents a 10-fold increase. The production of paclitaxel by the fungal strain was further improved via exposure to UV and gamma radiation at specific doses. The paclitaxel concentrations were intensified following UV and gamma radiation to 209·91 and 351·82 mg kg-1 . CONCLUSIONS: This is the first report on the production of paclitaxel using agro-industrial wastes as cheap source that may contribute in lowering the cost of producing paclitaxel. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings offer new and alternate sources with excellent biotechnological potential for paclitaxel production by fungal fermentation.

Biosynthesis of zinc oxide nanoparticles with antimicrobial, anticancer, antioxidant and photocatalytic activities by the endophytic Alternaria tenuissima.

AIMS: Zinc oxide nanoparticles (ZnONPs) were successfully synthesized using the culture filtrate of the endophytic fungus Alternaria tenuissima as a rapid, eco-friendly and cost-effective method. METHODS AND RESULTS: The rapid synthesis of ZnONPs was completed after 20 min as confirmed by UV-Vis spectroscopy. The synthesized ZnONPs showed a single-phase crystalline structure. Dynamic light scattering analysis showed that the synthesized ZnONPs were monodispersed and the recorded polydispersity index value was 0·311. Zeta potential value of -23·92 mV indicated the high stability of ZnONPs. Transmission electron microscope revealed the spherical shape and the mean particle size was 15.45 nm. Functional groups present in the prepared samples of ZnONPs were confirmed by Fourier transform infrared spectroscopy. Additionally, the biological activities of in vitro antimicrobial, anticancer, antioxidant as well as the photocatalytic activities were evaluated. ZnONPs showed broad spectrum of antimicrobial potential against all the tested plant and human pathogens. Based on the MTT assay, ZnONPs inhibited the proliferation of normal human melanocytes, human breast and liver cancer cell lines with IC50 concentrations of 55·76, 18·02 and 16·87 µg ml-1 . ZnONPs exhibited promising antioxidant potential with 50% inhibitory concentration of 102·13 µg ml-1 . Moreover, ZnONPs showed efficient degradation of methylene blue dye. CONCLUSIONS: The synthesized ZnONPs showed promising activities that can be better explored in the near future for many medical, agricultural and industrial applications. SIGNIFICANCE AND IMPACT OF THE STUDY: This study suggests a new and alternate approach with the excellent biotechnological potentiality for the production of ZnONPs which could open up the way for the industrial manufacture of nanoparticles using microbial platforms.

A novel source of the cardiac glycoside digoxin from the endophytic fungus Epicoccum nigrum: isolation, characterization, production enhancement by gamma irradiation mutagenesis and anticancer activity evaluation.

AIMS: Different endophytic fungi were isolated and screened for their digoxin-producing ability. Strain improvement and different culture conditions were studied for more effective production of digoxin. METHODS AND RESULTS: Among the isolated fungi, an isolate produced digoxin in a concentration of 2·07 mg l-1 . The digoxin-producing fungal isolate was identified as Epicoccum nigrum Link according to the morphological features and phylogenetic analyses. The potentiality of the fungal strain for production enhancement of digoxin was performed by gamma radiation mutagenesis. Gamma irradiation dose of 1000 Gy intensified the digoxin yield by five-fold. Using this dose, a stable mutant strain with improved digoxin productivity was isolated and the stability for digoxin production was followed up across four successive generations. In the effort to increase digoxin magnitude, selection of the proper cultivation medium, addition of some elicitors to the most proper medium and several physical fermentation conditions were tested. Fermentation process carried out in malt extract autolysate medium (pH 6·5) supplemented by methyl jasmonate and inoculated with 2 ml of 6-day-old culture and incubated at 25°C for 10 days stimulated the highest production of digoxin to attain 50·14 mg l-1 . Moreover, cytotoxicity of digoxin separated from the fungal culture was tested against five different cancer cell lines. Based on the MTT assay, digoxin inhibited the proliferation of the five different cancer cell lines and the recorded 50% inhibitory concentration ranged from 10·76 to 35·14 µg ml-1 . CONCLUSIONS: This is the first report on the production and enhancement of digoxin using fungal fermentation as a new and alternate source with high productivity. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings offer new and alternate sources with excellent biotechnological potential for digoxin production by fungal fermentation. Moreover, digoxin proved to be a promising anticancer agent whose anticancer potential should be assessed in prospective cancer therapy.

Immobilization technique for enhanced production of the immunosuppressant mycophenolic acid by ultraviolet and gamma-irradiated Penicillium roqueforti.

AIMS: Different entrapment matrices were screened to immobilize two strains of Penicillium roqueforti (AG101 and LG109) for more effective production of mycophenolic acid (MPA). Further improvement in the MPA productivity from immobilization of spores and mycelia was adopted by UV and gamma irradiation. METHODS AND RESULTS: Penicillium roqueforti strains were immobilized in different entrapping carriers and used for MPA production in shake flask cultures. Maximum MPA production was achieved on using an alginate concentration of 3·0% (w/v) and a mycelial fresh weight of 10% (w/v). MPA produced by alginate-immobilized spores and mycelia was almost double in comparison to the free system. The MPA-producing ability of immobilized AG101 and LG109 strain was significantly enhanced by mutagenesis through irradiation by UV (254 nm) for 120 and 90 min, respectively and gamma rays at 0·75 KGy. The feasibility of MPA production in a semi-continuous form by immobilized cells as affected by irradiation was adopted. CONCLUSIONS: MPA production by immobilized spores and mycelia was more intensified by UV and gamma irradiation. Moreover, the immobilized cell culture was superior to free-cell culture. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings indicate the future possibility to reduce the cost of producing fermentation-based drugs.