Anemarrhena asphodeloides Extract Inhibits the Mycelial Growth of Magnaporthe oryzae and Controls the Rice Blast Disease

Abstract

Previously, we have reported a plant extract isolated from Lysimachia foenum gracum Herba as a new environment friendly biopesticide that has the mycelial growth inhibition effect on Magnaporthe oryzae, the pathogenic fungus of the rice blast disease. For the finding of additional biopesticide candidate, we tested the mycelial growth inhibitory effects about 700 species of plant extracts on PDA media. Among them, the extract of Anemarrhena asphodeloides showed prominent inhibitory effect of which $IC_{50}$ was $139.7{\mu}g/ml$. Mycelial radii of M. oryzae were measured on PDA medium containing the four organic solvent fractions isolated from total extract from A. asphodeloides. Ethyl acetate fraction showed the impressive inhibitory effect of $IC_{50}$, $54.12{\mu}g/ml$. In the subsequent rice field test for the total extract of A. asphodeloides, we obtained encouraging 62.0% control rate of rice blast disease without any phytotoxicity. It is almost equivalent to that of chemical pesticides implying the applicability of the extract as a new biopesticide. In further study, the analysis of active ingredients of the extract would be necessary for the development of a new biopesticide and for the verification of cellular mechanism by which the mycelial growth of M. oryzae inhibited.

Fig. 1. Massive screening for the mycelial growth inhibitory effects of 700 kinds of plant extracts on M. oryzae. Plant extracts provided by plant extract bank (Korea) were tested on PDA solid media in 24 well plates by empirical concentration of 500 ㎍/㎖ of each plant extracts.

Fig. 2. Mycelial growth inhibitory effect of extract of A. asphodeloides on M. oryzae in dose dependent treatment. (A) None treatment. Extract was treated by the concentrations of 10, 50, 100, 150, 200, 300, 400, 500, 600, and 700 ㎍/㎖, respectively. Mycelia of M. oryzae cut by cork borer were incubated at 28℃ on the PDA solid media for 5 days. (B) The diameters or radii were determined by mean value of individual measured values. Those experiments were performed in triplicate (p<0.05, Anova test).

Fig. 3. Fractionation of methanol total extract of A. Asphodeloides extract. Each fraction was prepared by orderly treatment of n-Hexan, chloroform, ethyl acetate and n-butanol and were vaporized for the treatment of mycelial growth inhibitory test.

Fig. 4. Mycelial growth inhibitory effect of fraction. (A) Hexane, Chloroform, Ethyl acetate, and n-butanol fraction were tested by the concentration of 200 ㎍/㎖ of each fraction. (B) Mycelial radii of M. oryzae were measured on the PDA solid media for 5 days after the treatment of ethyl acetate fraction. Those experiments were performed in triplicate (p<0.05, Anova test).

Fig. 5. View of test port of rice blast disease field test of rice plant, Chucheng. (A) whole view of testport of none treatment, (B) Symptoms of rice blast disease in none treatment test port by M. oryzae, (C) Whole view of treatment testport, (D) Test port of A. asphodeloides extract treatment. Those experiments were performed in triplicate.

Table 1. Effect of A. asphodeloides extract on control rate of rice in field test

Table 2. Effect of A. asphodeloides extract on phytotoxicity of rice in field test