Effects Of Three Stilbene Compounds On The Growth Of Aspergillus Niger And The Production Of Ochratoxin A (OTA)

Ochratoxin A(OTA)is a fungal secondary metabolite with toxicity and one of the most harmful mycotoxins to human beings.The pollution of OTA to grape and its products is very serious all over the world,which causes great economic loss.Aspergillus carbonarius(A.carbonarius)is the main fungus in the production of OTA.Therefore,it is an important measure to prevent the OTA pollution in grape and its products by inhibiting the growth and toxin production of A.carbonarius.Stilbene is an important natural polyphenol compound,which has many biological activities such as anti-inflammatory,anti-pathogen and anti-oxidation.Resveratrol,piceatannol and pterostilbene were used as antibacterial agents to study their effects on the growth of A.carbonarius and the production of OTA in vitro.In addition,the inhibitory effect of pterostilbene,which has the strongest antifungal activity in vitro,on the growth of A.carbonarius and the production of OTA in grape juice and the maintenance of nutritional quality were explored.The main results are as follows:1.The antifungal activity of stilbenes against A.carbonarius showed that the inhibition effect of stilbenes on the mycelial growth and spore germination of A.carbonarius was from strong to weak:pterostilbene>piceatannol>resveratrol,the IC50 values of resveratrol,piceatannol and pterostilbene were 5.1mM,1.8mM and 0.28mM,respectively.It was found by SEM and TEM that the treatment of pterostilbene lead to the incomplete sporangium,the bending and breaking of mycelial,the breaking of cell membrane and the aggregation of contents.With the increase of culture days,the degree of grape decay increased.The content of OTA and stilbenes in grape were detected by UHPLC-QQQ-MS/MS,and the results showed that the content of OTA and stilbenes in grape by infected A.carbonarius were increased obviously.At the early stage of infection(48 h),stilbenes could control the synthesis of OTA(48 h),but with the increase of culture days,the synthesis of OTA was less affected by stilbenes.2.The experimental results of the influence of stilbene compounds with different concentrations on the production of OTA are as follows:when the concentration was lower than 0.2mM,the OTA production in treatment group of resveratrol and piceatannol was significantly higher than that in the blank group,but the OTA production could be inhibited at 0.4mM.The inhibitory effect of pterostilbene on OTA production increased with the increase of concentration.The OTA production increased first and then decreased with the change of culture days,and the production was the most on the sixth day.The production of OTA was about 5ng/mL under the treatment of stilbenes and different time points at shaking culture,and it was 350-700 times less than that of static culture.The trend of OTB was consistent with that of OTA in all experimental groups.There was no correlation between the mycelial dry weight of A.carbonarius and OTA production,that is to say,higher mycelial dry weight did not necessarily produce higher OTA.RT-qPCR results showed that the most up and down regulation changes of PKS?NRPS?HAL?P450?bZIP and LaeA genes related to OTA biosynthesis were consistent with the change of OTA content.Some of the inconsistencies may be caused by the effect of transcription or translation.3.Because OTA pollution mainly occurs in grapes and grape products,and the above results show that pterostilbene has the strongest inhibitory effect on the growth of A.carbonarius and OTA production in vitro.Therefore,the inhibitory effect of pterostilbene on A.carbonarius and the protective effect on nutritional quality in grape juice were studied.The results showed that pterostilbene could inhibit the growth of A.carbonarius and the production of OTA in grape juice,and the content of OTA increased with the increase of culture days,and the content of OTA increased sharply on the 9th day.When the grape juice was infected by A,carbonarius,the content of reducing sugar and soluble solids and pH value in grape juice decreased with the increase of culture days,and titrated acid increased significantly.In addition,the infection of A.carbonarius could also lead to the raise of the content of tartaric acid,citric acid and malic acid content,and the decrease of the content glucose and fructose,but the treatment of pterostilbene could make all the above indexes more close to the juice not infected by A.carbonarius.