Inhibition of Ethylene Production in Fruit Slices by a Rhizobitoxine Analog and Free Radical Scavengers

Abstract

The rhizobitoxine analog, L-2-amino-4-(2-aminoethoxy)-traas-3-butenoic acid (Ro), wich effectively Iits ethylene productio in apple (Mahus domestca Brkh.) and otber tissues at concentratins at about 68 micromolar, inhibited ethylene productin by about 50 to 70% in green tomato (Lycopersicon esculentum Mill.) Since Lieberman et al. (6) showed that ethylene was derived from carbon atoms 3 and 4 of methionine in apple slices, evidence has accumulated that methionine is the natural precursor of ethylene in several plant tissues (2,11). Despite this evidence, the exact reaction mechanism of the conversion of methionine to ethylene has not been established. With the discovery that rhizobitoxine inhibits ethylene production from methionine (1 1), a tool was provided for probing the pathway(s) of ethylene biosynthesis in plant tissues. Rhizobitoxine was found to inhibit effectively ethylene production of apple and banana slices, pea seedlings, tobacco leaves, sorghum seedlings (11), and flower tissues (5); the assumption was that methionine was the major precursor of ethylene in those tissues. Rhizobitoxine and certain of its analogs (Ro)' inhibit ethylene production irreversibly, presumably by binding to an enzyme involved in the first step of the methionine to ethylene pathway (7). One purpose of the present investigation was to extend study of the rhizobitoxine effect of other fruits and to determine whether sensitivity of the ethylene-producing system to rhizobitoxine varies with the different stages of maturation and senescence. A second purpose was to determine whether free radical scavengers inhibit ethylene production in vivo, as they have been shown to do in a model system which converts methional to ethylene following hydroxyl radical attack (4). A preliminary report of this work was published earlier (1).' Abbreviations: Ro: ethoxy analog of rhizobitoxine; Inhib.: inhibition. MATERIALS AND METHODSPlant Material and Procedure. Tomato (Lycopersicon esculentum Mill., cvs. Rutgers, Cal Ace, and Walter) and apple (Malus domestica Borkh., Red Delicious) fruit were grown at the Beltsville Agricultural Research Center, and unknown varieties of tomato were purchased from a local wholesale market. The responses of tomato fruit slices to inhibitors of ethylene production appeared to be independent of variety. Avocado (Persea gratissima Gaertn.) fruit were from a local market or provided by Calavo Growers of California. Tomato pericarp tissue was cut into about 0.5-cm cubes; avocado and apple tissues were cut into discs about 0.3 cm thick and I cm in diameter. Tissue samples (2 g) were incubated in 25-ml stoppered Erlenmeyer flasks containing 3 ml of 0.4 M sucrose in 0.027 M citrate-0.047 M Na-phosphate buffer (pH 4.6).The rhizobitoxine analog, sodium benzoate, gailic acid, and propyl gallate stock solutions were made up in that buffer. Nordihydroguaiaretic acid and eugenol solutions were prepared with ethanol, and the final concentration of ethanol in the incubation flasks was 1.5%. The flasks were incubated ...

Bicarbonate/CO2-Facilitated Conversion of 1-Amino-cyclopropane-1-carboxylic Acid to Ethylene in Model Systems and Intact Tissues

“…Several lines of evidence from both intact tissues and model systems suggest that ethylene formation from ACC may be mediated by free radicals. For example, the reaction can be inhibited in situ by radical scavengers (4,5), is driven by OH in a strictly chemical system consisting of ACC and the components of the Fenton reaction (22), and appears to be facilitated by 02 when catalyzed by isolated microsomal membranes (25). Thus, it is conceivable that bicarbonate might enhance ACCdependent ethylene production by facilitating the formation of these reactive species of oxygen.…”

Methionine-induced Ethylene Production by Penicillium digitatum

“…Propyl gallate, which quenches free radicals and thus inhibits ethylene production in higher plants (1) and in the horseradish peroxidase model system (25), was as effective an inhibitor of ethylene evolution as boiling. Chelators which have been shown to inhibit ethylene production from methionine in both the model systems (14,17) and in apple slices (15) also somewhat inhibited ethylene production by the incubating solution.…”

“…The difference between these results may be due to a 20-fold difference in concentration of the inhibitors used. Although the effects of rhizobitoxine and its analogue on ethylene production by higher plants are similar (1,16,19), their effects may be different in the fungus and may be related to inhibition of transaminase, which converts a-ketoglutarate to glutamate (21). In shake cultures RO only slightly inhibited the production of ethylene stimulated by methionine (Table I).…”

Inhibition of Ethylene Biosynthesis by Salicylic Acid

“…LESLIE AND ROMANI Baker et al (2) demonstrated inhibition of ethylene biosynthesis by benzoic acid at concentrations that were also active in this study (Table I), but benzoic acid was considerably less active in pear cells than SA or ASA. This disparity in ability to block ethylene biosynthesis contrasts sharply with observations ofbenzoic acid activity equal to or stronger than that of SA in flower induction (10,26), vegetative growth (12), and disease resistance (27), suggesting different mechanisms may be involved.…”