Development Of H₂O₂ Electrochmical Sensor And Its Application For Studying Of The Mechanism Of Aflatoxin Inhibition In Aspergillus Flavus By Citral

Many researches have showed that citral can induce decrease of aflatoxin production from Aspergillus flavus effectively. But the inhibition mechanism of citral to aflatoxin production is not clear.A novel non-enzymatic electrochemical H2O2 sensor was fabricated. The citral-triggered H2O2 secretion from A. flavus cells was real-time monitored with the sensor to study the inhibition mechanism of citral to aflatoxin production, which could provide a theoretical basis for the prevention of aflatoxin contamination in the food. The main conclusions are listed as follows:1. PDA-Fe3O4 nanocomposites were prepared via an in situ co-precipitation approach. SEM and TEM data illustrate that Fe3O4 NPs are successfully synthesized and coated on the surface of the PDA microspheres uniformly. The nanocomposites have good hydrophilicity estimated by the FTIR spectra.2. A novel non-enzymatic H2O2 non-enzyme sensor was fabricated by dripping PDA-Fe3O4 nanocomposites on GCE. The sensor exhibits good selectivity, a linear dynamic range of 0.5 to 6.0μM, a sensitivity of 51.06 μA/m M and a detection limit of 4.9×10-8 M(S/N = 3). And it was applyed in situ H2O2 detection in biological systems successfully.3. Detected the concentration of aflatoxin in different culture time with different treatments by HPLC.The results showed that aflatoxin production can be inhibited by both short-time and long-time treatment with citral.4. The ROS level was analyzed with an electrochmical sensor and a fluorescence-based approach.The accumulation of AFB1 in the culture solution was measured with HPLC. The activities of anti-oxidative enzymes including CAT, SOD and GSH-Px, as well as the content of MDA and protein carbonyl, were also monitored to indirectly reflect the intracellular ROS level. The surface morphologies of A. flavus cells were characterized with scanning electron micrographs(SEM) to investigate the effects of citral treatment on the cell membrane. The results indicated that the inhibition mechanism of citral to aflatoxin production has close relationship with ROS, and the mechanism may be include short-term effect and long-term effect. Citral triggered ROS secretion from A. flavus cells just seconds after being added, which make the concentration of intracellular ROS can’t meet the level aflatoxin production need, inhibiting aflatoxin production. Citral can induce A. flavus generate ROS for a long time, making the the concentration of ROS beyond the cearing ability of antioxidant enzyme. The excess ROS can cause oxidative damage to A. flavus,making it unable maintain normal physiological condition and product aflatoxin.