Study On Antifungal Effect And Synergistic Mechanism Of Eugenol-nanoethosomes

Eugenol is a secondary metabolite in plants.It is a volatile compound at room temperature with a aroma.Eugenol is generally recognized as a safe substance which can be used in food.In particular,its effective antifungal activity is potential in the application of fruit and vegetable preservation.Nanoethosome is a kind of nanoparticles with special structure,which possesses surface effect and quantum size effect.Nanoethosome can be used to load eugenol to prepare eugenol-nanoethosome.The nanoethosome is potential to improve the antifungal effects of eugenol in fruit and vegetable preservation due to the excellent permeability and sustained-release of nanoethosomes.The present study aims the preparation,antifungal improvement of eugenol-nanoethosome.(1)The optimum conditions for preparing eugenol-nanoethosome are studied.The results showed that the optimum eugenol-nanoethosome compsoed of eugenol 0.5%(v/v),phospholipid 20%(v/v)and ethanol 30%(v/v).Eugenol-nanoethosome with the highest antifungal activity showd average diameter 44.21 nm,PDI 0.189,and zeta potential-40.3m V.(2)The minimum inhibitory concentration(MIC)of eugenol-nanoethosome on mycelium was 95 u L/L and 79 u L/L respectively.Eugenol-nanoethosome exhibit higher MIC due to the slow-release effect of nanoethosomes.Furthermore,eugenolnanoethosome showed higher mycelial inhibition rate than non-nanoethosome by 5.17%,3.93%,6.1% and 2.75% for the anthracnose from banana,strawberry,grape and loquat,respectively.The spore inhibition rate of eugenol-nanoethosome was 11.52%,7.82%,2.54% and 10.47% for the four fungi,which higher than that of non-nanoethosome,respectively.The antifungal activity of eugenol-nanoethosome on fruit loquat in vivo was further verified.After inoculation of 6 days,the incidence of anthracnose in control group and non-nanoethosome was 100%,while that of eugenol-nanoethosome was 80%.The diameter of lesions in eugenol-nanoethosome treatment group was smaller than that in non-nanoethosome treatment group.The inhibiting rate of 5 ?L/L non-nanoethosome fumigation was 52.24%,while that of eugenol-nanoethosome was 85.75%.(3)The results of fluorescent dye tracking experiments showed that the penetration depth of the nanoethosome in epidermis was deeper than that of nanoethosome and free eugenol groups.The content of eugenol in different parts of epidermis was detected by GC-MS.the content of eugenol in inner epidermis and nanoethosome group was 31.3%and 91.42% higher than that in non-nanoethosomes group and eugenol group.The results show that nanoethosome have good permeability and antifungall activity by delivering eugenol into tissues.The results showed that the inhibition rate of non-nanoethosome was4.5%,while that of nanoethosome was 62.35%,which increased by 57.85%.Therefore,the nanoethosome has good permeability,and the eugenol can reach the parasite site(inner surface layer)of the pathogen to achieve antifungall effect.(4)The study of sustained release of nanoethosome showed that nanoethosome could effectively improve the antifungal effect of eugenol against Colletotrichum gloeosporioides.Free eugenol,non-nanoethosome and eugenol-nanoethosome with the same content of eugenol were exposed to air for 0-10 h,and then were used to inhibit the growth of mycelium.The results showed that the eugenol completely lost the antifungal effect after 4 h volatilization of free eugenol and non-nanoethosome,while eugenol-nanoethosome were exposed to air for 8 h,the antifungal efficiency was still more than 40.84%.The inhibition of spore growth further showed that the nanoethosome showed better antifungal effect,and the inhibition effect on spore growth was up to 42 h,and the inhibition time was effectively prolonged by 18 h and 13 h compared with free eugenol group and non-nanoethosome group,respectively.After 12 h,the content of eugenol in the medium was higher than those of the non-nanoethosome group and free eugeno group,which indicated that eugenol-nanoethosome had good sustained-release.Through the above contents,the antifungall and synergistic effect and mechanism of nanoethosome was preliminarily clarified,and the mechanism of its increasing inhibition effect was further studied,which can provide reference and technical basis for the application of plant essential oil in fruit and vegetable preservation.