Azoxystrobin assay

The detached leaf assays indicated that 625 ppm CAY-1 and sampangine provided effective protectant activity for disease control of anthracnose on the leaf surface (Table 1). These compounds were as effective as the commercial fungicide, azoxystrobin. 

Experimental compounds were evaluated in a dose-response format. Azoxystrobin or other fungicides that have both protective and curative activity were used as a standard, and a solvent control was used in every study. The number of disease lesions per leaf was used to determine the ability of the test compounds to prevent infections. The size of the lesions was used to determine the curative activity of compounds. Each fungicide concentration is replicated four times and the experiment is repeated at least once. This bioassay does not differentiate between direct effects on the fungus and indirect effects through induction of plant defenses. However, if a compound is much more active in this in vivo assay than than the in vitro microtiter assay, induction activity is indicated. 

Detached leaf assays provide us with the opportunity to evaluate new fungicides directly on the leaf surface in a dose-response format (Table 1). This assay allowed us to benchmark potential lead compounds such as CAY-1 and sampangine with a commercial standard (azoxystrobin) of known mode of action (Qo I inhibitor). The number of diseased lesions was used to determine effective concentrations needed for disease control. Lesion size is used to determine the relative effectiveness of the systemic activity that produced curative activity 24 hrs after inoculation. The detached leaf assay was also used to establish experimental field rates for future studies. Study of protectant activity indicated that 1250 ppm. CAY-1 or sampangine appeared to be an effective concentration for disease control of anthracnose on the leaf surface, or between 100-1000 times the concentration required for in vitro activity (Post, Table 1). 

The media controls (medium without added cells) played two roles, providing information on those compounds which are not stable in aqueous conditions, and also determining if the compound under test was soluble under aqueous conditions. As shown in Figure 3, the level of pyraclostrobin in the incubations showed a decline over a 72 hr period. In this case, no breakdown products were observed and it is likely that the pyraclostrobin precipitated out of solution. Azoxystrobin, on the other hand, remained in solution, although there was some variation in the amounts observed at the different time points indicating that there was some variability in the assay. Due to the qualitative nature of this assay, a high level of precision is not required. None of the other compounds tested were either unstable or insoluble under the incubation conditions. 

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