Insecticidal okaramines

As mentioned earlier, P. simplicissimum ATCC 90288 produced insecticidal okaramines. Moreover, this strain induced the same effect on the silkworms as verruculogen (83). The acetone extract obtained from the mycelia and media of this strain was concentrated and the aqueous residue was extracted with dichloromethane. The dichloromethane extract was partitioned between hexane and methanol, containing 10% water. The activity was found only in the lower layer. The active ethyl acetate extract obtained from the lower layer was chromatographed on silica gel with a hexane-ethyl acetate mixture. The 40-60% ethyl acetate eluates were rechromatographed on silica gel with a hexane-chloroform mixture. HPLC of the active 90-100% chloroform eluates on a Capcell pack Ci8 column, using 65.7% aqueous methanol with a flow rate of 1.0 ml/min, yielded two active compounds, AC 1 and AC 2. 

The material from an acetone extract of okara fermented with P. simplicissimum ATCC 90288 was partitioned between ethyl acetate and water. The ethyl acetate layer thus obtained exhibited insecticidal activity. The ethyl acetate fraction was repeatedly chromatographed on silica gel with a hexane-acetone mixture, and then on alumina with a hexane-ethyl acetate mixture. Crystallization of the 50% ethyl acetate eluate and of 60-80% ethyl acetate eluates gave two active compounds, which were named okaramines A (1) and B (2), respectively [13-16],... 

From an okara medium fermented with P. simplicissimum AHU 8402, three insecticidal compounds were isolated. Two of them were identified as 1 and 2, and the third one seemed to be a new, related compound, which was thus named okaramine C (4). Okaramine C (4), C32H36N4O3, proved to be a tetrahydro-derivative of 1. The -NMR and 1H-H COSY spectra indicated that 4 had an additional reverse-prenyl group, an exchangeable proton and a -CH2-CH< group instead of -CH=CH- and -CH=C<, suggesting that the C-r=C-2 double bond was saturated and... 

In further screening microbes for insecticides, an isolate Aspergillus aculeatus Iizuka KF-428 was obtained from a soil sample. This strain also exhibited the insecticidal activity when grown on okara, and three active principles were isolated. Two of them were identified as okaramines A (1) and B (2). The third one also seemed to be an okaramine-related compound and was named okaramine H (9) [20]. 

Table 1. Insecticidal Activity of Okaramines against Silkworms.

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