Ma Weibin, Zhao Luling, Johnson Eric T, Xie Yanli, Zhang Mingming
Department of Pharmacy, Henan University of Chinese Medicine, Zhengzhou 450046, China; Henan Key Laboratory of Cereal and Oil Food Safety Inspection and Control, Henan University of Technology, Zhengzhou 450001, China.
Henan Key Laboratory of Cereal and Oil Food Safety Inspection and Control, Henan University of Technology, Zhengzhou 450001, China.
Int J Food Microbiol. 2022 Jun 2;370:109633. doi: 10.1016/j.ijfoodmicro.2022.109633. Epub 2022 Mar 15.
Natural food flavour (E)-2-hexenal, a green leaf volatile, exhibits potent antifungal activity on Aspergillus flavus, but its antifungal mechanism has not been fully elucidated. In this study, we evaluated (E)-2-hexenal-induced apoptosis in A. flavus conidia and explored the underlying mechanisms of action. Evidence of apoptosis in A. flavus conidia were investigated by methods including fluorescent staining, flow cytometry, confocal laser scanning microscope, and spectral analysis. Results indicated that 4.0 μL/mL (minimum fungicidal concentration, MFC) of (E)-2-hexenal application induced early markers of apoptotic cell death in A. flavus conidia with a rate of 38.4% after 6 h exposure. Meanwhile, typical hallmarks of apoptosis, such as decreased mitochondrial membrane potential (MMP), activated metacaspase activity, fragmented DNA, mitochondrial permeability transition pore (MPTP) opening and cytochrome c (Cyt C) release from mitochondria to the cytosol were also confirmed. Furthermore, intracellular ATP levels were reduced by 63.3 ± 3.6% and reactive oxygen species (ROS) positive cells increased by 31.1 ± 3.1% during A. flavus apoptosis induced by (E)-2-hexenal. l-Cysteine (Cys), an antioxidant, could strongly block the excess ROS generation caused by (E)-2-hexenal, which correspondingly resulted in a significant inhibition of MPTP opening and decrease of apoptosis in A. flavus, indicating that ROS palys a pivotal role in (E)-2-hexenal-induced apoptosis. These results suggest that (E)-2-hexenal exerts its antifungal effect on A. flavus conidia via a ROS-dependent mitochondrial apoptotic pathway.
天然食品香料（E）-2-己烯醛是一种绿叶挥发物，对黄曲霉具有强大的抗真菌活性，但其抗真菌机制尚未完全阐明。在本研究中，我们评估了（E）-2-己烯醛诱导黄曲霉分生孢子凋亡的情况，并探讨了其潜在的作用机制。通过荧光染色、流式细胞术、共聚焦激光扫描显微镜和光谱分析等方法研究了黄曲霉分生孢子凋亡的证据。结果表明，施加4.0μL/mL（最低杀菌浓度，MFC）的（E）-2-己烯醛可诱导黄曲霉分生孢子出现凋亡细胞死亡的早期标志物，暴露6小时后凋亡率为38.4%。同时，还证实了凋亡的典型特征，如线粒体膜电位（MMP）降低、metacaspase活性激活、DNA片段化、线粒体通透性转换孔（MPTP）开放以及细胞色素c（Cyt C）从线粒体释放到细胞质中。此外，在（E）-2-己烯醛诱导黄曲霉凋亡过程中，细胞内ATP水平降低了63.3±3.6%，活性氧（ROS）阳性细胞增加了31.1±3.1%。抗氧化剂L-半胱氨酸（Cys）可强烈阻断（E）-2-己烯醛引起的过量ROS生成，这相应地导致黄曲霉中MPTP开放的显著抑制和凋亡的减少，表明ROS在（E）-2-己烯醛诱导的凋亡中起关键作用。这些结果表明，（E）-2-己烯醛通过ROS依赖性线粒体凋亡途径对黄曲霉分生孢子发挥抗真菌作用。
