In this study, 3-nitropropionic acid (3-NPA) was separated and purified from endophytic fungi belonging to Phomopsis sp. and its cytotoxicity was determined by MTT assay. Treatment with 3-NPA for 24 h resulted in a dose-dependent apoptosis in MCF-7 cells. Through quantitative detection of the genes that are closely related to the Bcl-2 signalling pathway, there was an increased expression of p53 and Bax and a decreased expression of Bcl-2, which indicated apoptosis in these cells. Meanwhile, the overexpression of PARA (poly ADP-ribose polymerase) and apoptosis inducing factor (AIF) also suggested that 3-NPA induced cellular apoptosis through a caspase-3-independent pathway in caspase-3-deficient MCF-7 cells. The fermentation condition was also improved to produce more 3-NPA: glucose as a carbon source and yeast extract as a nitrogen source, fermentation for 8 days at 32°C and a solution environment of pH 5·0. Under these conditions, the yield of 3-NPA was increased to 529 mg l(-1) compared with 410 mg l(-1) under traditional fermentation conditions.
Significance and impact of the study: 3-Nitropropionic acid is a mitochondrial inhibitor and has some useful bioactivities such as antibacterial activity. In this paper we found that 3-NPA also has obvious cytotoxicity, so we studied its antitumour activity and tried to determine the antitumour molecular mechanism, opening a new perspective for potential antitumour prodrug development. As 3-NPA is often obtained from natural products with a low yield, in order to overcome the disadvantage of an endophytic fungi source of 3-NPA, we optimized the fermentation conditions for 3-NPA in Phomopsis sp. to obtain the maximum production of 3-NPA.
Keywords: 3-nitropropionic acid; Bcl-2 signalling pathway; antitumour activity; apoptosis; caspase-3-independent; fermentation condition optimization.
© 2015 The Society for Applied Microbiology.