Plant growth regulator diethyl aminoethyl hexanoate suppresses Aspergillus flavus growth and aflatoxin biosynthesis through disruption of redox homeostasis

ABSTRACT Aflatoxin B₁ (AFB₁), produced by Aspergillus flavus , is a major threat to food safety and human health. In this study, we report that diethyl aminoethyl hexanoate (DA-6), a plant growth regulator, exhibits strong antifungal and anti-aflatoxigenic activities against A. flavus . DA-6 significantly inhibited mycelial growth, conidial germination, and sclerotia formation in a dose-dependent manner, while also effectively suppressing fungal colonization in stored rice bran and maize. Microscopic and biochemical analyses revealed that DA-6 disrupted plasma membrane integrity, leading to cytoplasmic leakage and hyphal collapse. Further analyses demonstrated that DA-6 caused severe oxidative-redox imbalance, characterized by excessive reactive oxygen species (ROS) accumulation and impaired antioxidant defenses, which were accompanied by mitochondrial dysfunction, reduced membrane potential, and ATP depletion. Transcriptomic profiling confirmed a global repression of the aflatoxin biosynthetic gene cluster (including aflR and aflS ) and key developmental regulators ( laeA and veA ), resulting in the near-complete cessation of AFB₁ production. Additionally, functional characterization of two highly responsive Zn₂Cys₆ transcription factors (AFLA_059960 and AFLA_084720) further supported the involvement of oxidative stress adaptation pathways in the fungal response to DA-6-induced stress. Collectively, these findings provide mechanistic insights into the antifungal action of DA-6 and highlight its potential application for controlling fungal contamination and aflatoxin risks in agricultural commodities. IMPORTANCE Plant growth regulators (PGRs) are widely used in agriculture, yet their potential effects on fungal physiology and mycotoxin biosynthesis remain largely unexplored. In this study, we demonstrate that diethyl aminoethyl hexanoate (DA-6) caused extensive transcriptional repression of fungal developmental regulators and the aflatoxin biosynthetic gene cluster, accompanied by severe oxidative-redox imbalance and mitochondrial dysfunction. These findings reveal a previously unrecognized antifungal application of PGRs and provide new insights into their potential use for controlling toxigenic fungi and mycotoxin contamination in food and agricultural systems.