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Alternaria blotch of apple, caused by a pathogenic strain of Alternaria mali Roberts, has been one of the destructive diseases since 1956 in Japan. The apple cultivars Indo and Delicious are particularly susceptible and Jonathan is resistant1). One of the authors previously reported that the culture filtrate of the fungus caused necrotic brown spots on leaves of susceptible cultivars as the pathogen did and therefore the fungus might excrete a host-specific toxin. However, the toxin was unable to isolate as a highly purified state2). So far host-specific toxins which were isolated as a chemically pure compound have been known only in a few cases3). In this paper we wish to report the isolation of the host-specific toxin produced by the title fungus. Alternaria mali Roberts (AKI-3 strain) was grown in the Richards' medium in stationary culture for 2-3 weeks at 37C. The culture broth (pH 5.5-6.5) thus obtained was filtered and the fitrate was repeatedly extracted with chloroform, and the extract was concentrated to give a reddish oily material in vacuo. The aqueous solution was treated with active charcoal and the charcoal was separated. The dried charcoal was eluted with acetone and then the acetone solution was evaporated in vacuo to give a reddish oil. The host-specific toxicity was examined by the biological test, whether or not necrotic brown spots could be formed when a few drops of aqueous solution of the extracts put on green leaves of Indo respectively. As a result of this test it was found that the chloroform extract showed much more potent toxicity than the acetone eluent. On the other hand, the chloroform extract of the frozen mycelium also exhidited strong toxicity and the chromatographic behaviour of this extract was identical with that of the filtrate. Thereafter the chloroform extract was chromatographed on silica gel and elution with chloroform-ethyl alcohol (93:7v/v) gave crude crystals. Recrystallization from chloroform-ethyl alcohol afforded pure crystals, mp 220-222C, which showed only one spot on silica gel thin layer chromatography (chloroform-ethyl alcohol, 95:5v/v). The toxicity of the pure compound was again examined and the aqueous solution of such a low concentration, 1ƒÊg/ml, caused brown spots on leaves of the susceptible cultivar, Indo. Contrarily on leaves of Jonathan the solution of the higher concentration, 1mg/ml, did not cause any changes (Fig. 1). Consequently, it is evident that a host-specific toxin produced by the fungus was isolated. In the mass spectrum of this compound the peak at m/e 445.2194 is considered to be the molecular ion

Isolation of a Host-specific Toxin Produced by Alternaria mali Roberts