Preliminary Study On Antifungal Activity And Mechanism Of Carvacrol Aganist Two Important Plant Pathogenic Fungi

The rice blast and grey mold disease caused by Magnaprothe oryzae and Botrytiscinerea are all fungal disease which have great dangers and cause a heavy loss toagricultural production. So far they mainly rely on chemical control, but chemical controlcan cause environmental pollution and make the fungi resistant which weaken controleffect highly. Therefor, for sustainable development and the need of green chemistry,botanical fungicide are getting more and more attention, they not only do not causeenvironmental pollution，but also do not make the fungi resistant.Carvacrol is one of the main components of Crofton Weed volatiles. It had beenreported to be a broad-spectrum antifungal agent and antibacteria agent, and non-toxic、non-allergenic. However, there are no report on the effect of carvacrol on Magnaprotheoryzae and Botrytis cinerea and its mechanism of action. In this paper, antifungal activityof carvacrol against Magnaprothe oryzae and Botrytis cinerea and its mechanism of actionwere studies in vitro. The results were described as the follwing：（1）Antifungal activity of carvacrol was determined by mycelial growth rate andspore germination method, it was found that carvacrol had a strong inhibitory effect onMagnaprothe oryzae and Botrytis cinerea and its inhibitory medium concentration（EC50）were36.07mg/L and10.65mg/L directly by experiment determination. By the treatmentwith the carvacrol, the spore germination of Magnaprothe oryzae and Botrytis cinereashowed no regularity and prominence comparing with CK.（2）Light and SEM, TEM observation showed that by the treatment with the carvacrol, the mycelial morphology of Magnaprothe oryzae and Botrytis cinerea showedabnormality. For Magnaprothe oryzae，when carvacrol concentration is equal or greaterthan7.5mg/L, the mycelium showed twist、uneven thickness、cytoplasm pyknosis; whencarvacrol concentration is equal or greater than30mg/L, mycelium top and local inflation;when carvacrol concentration is equal or greater than60mg/L, increased formation ofbranches、hyphal vacuoles、unclear cell walls and broken mycelium were observed. ForBotrytis cinerea, when carvacrol concentration is equal or greater than3.125mg/L, themycelium showed cytoplasm pyknosis and appeared vacuoles; when carvacrolconcentration is equal or greater than6.25mg/L, the mycelium showed twist and uneventhickness; when carvacrol concentration is equal or greater than12.5mg/L, increasedformation of branches、tip defects、local inflation、broken mycelium were observed; whencarvacrol concentration is equal25mg/L, mycelium breakage、cavity and deformity wereserious，cell walls were unclear completely.（3）After adding carvacrol, the penetrability of cell membrane changed, resulting inleakage of inclusion of mycelium, for specific performance the conductivity of the culturemedium showed a increasing trend as the increases of concentration and time. The contentof soluble protein in mycelium was determined with Coomassie Brilliant Blue G-250colorimetric method. By treatment with carvacrol, the content of soluble protein wasdeclined.（4）We found that the chitinase activity and the N-acetyl glucosamine contentincreased first and then decreased with the prolonging of treatment time after addingcarvacrol by experiments on chitinase activity and N-acetyl glucosamine content assay.（5）Ergosterol was extracted by the saponification method, then measured by HPLC,the results showed that with the increase of the concentration, the content of ergosteroldecreased showing that the membrane structure was damaged.（6）Through experiments about the catalase activity assay, the catalase activitydecreased resulting in reduction of detoxification.The cell of fungi were death.