tion of avocado (Persea americana) or apple (Pyrus nal/us) fruit plugs. When the color of the Shamouti peel was orange, the ability to produce ethylene and to ...
Plant Physiol. (1970) 45, 533-534
Short Communication
Ethylene Production by Citrus Fruit Peel STIMULATION BY PHENOL DERIVATIVESI1 2 Received for publication August 28, 1969
YORAM FUCHS The Volcani Institute of Agricultural Research, Rehovot, Israel There is still no complete agreement concerning the role of ethylene in the natural ripening of citrus fruits (8), but it has been shown that under stress conditions (8) and at a certain fruit age (1) ethylene is evolved by citrus fruit. This implies that there may be an ethylene-producing system in citrus fruit which is mostly inhibited or simply not activated under normal ripening conditions. It has already been shown that phenolic compounds are required in the enzymatic conversion (model system) of methional to ethylene (10). In studies of the enzymatic conversion in vitro of methional to ethylene, p-coumaric acid was required as a cofactor (5, 7), and a strong inhibitory effect of catechol (5, 10) and resorcinol (5) on the ethylene synthesis was
to 6 mm thick, were placed in a 50-ml flask, with 2 ml of a test solution. All plugs were placed so that the albedo was facing the bottom of the flask, and the flavedo remained uncovered by the solution. The flasks were sealed with one-hole stoppers with clamped capillary tubes; the atmosphere in the flasks was sampled after 20 hr at 20 C and tested for ethylene by gas chromatography with a flame ionization detector and alumina column. Table 1. Effect of Some Phenzolic Compountds otn Ethylente Productionz by Plugs of Citrus Fruit Peel antd of Avocado anzd Apple Fruit All compounds were dissolved in 0.1 M phosphate buffer, pH 7.0, at 103 M. Addition
Tissue Tested
Ethylene Production
(plgfresh wi- 20
hr) X 104
Shamouti peel, green
pH
FIG. 1. The effect of pH of the incubating medium on ethylene production. Five plugs of green peel of Shamouti orange fruit were incubated in 2 ml of different buffer solutions (0.1 M) in 50-ml sealed flasks. Acetate buffer, pH 3 to 4.5; phosphate buffer, pH 5.5 to 7.6; and sodium bicarbonate, pH 8.4. Ethylene was determined after 20 hr of incubation at 20 C, and data are averages of three experiments.
Catechol Catechol + 2,3naphthalenediol 2, 3-Naphthalenediol 4-Methyl catechol p-Cresol Guaicol p-Coumaric acid Resorcinol
Quinol None
Shamouti peel, orange Clementine peel, green Avocado fruit plugs
reported. In a recent study on abscission (2) it was concluded that catechol (10'6 M) did not accelerate the abscission of coleus explant petioles, but it doubled the ethylene production at the nodes. Catechol (1000-2000 ,ug/ml) was also tried unsuccessfully as a fruit abscission-inducing agent on citrus trees (9). In our study of the ethylene production of Shamouti orange (Citrus sinensis L.) peel, catechol and 4-methyl-catechol showed a similar stimulatory effect. Five Shamouti orange peel plugs, 15 mm in diameter and 5
Apple fruit plugs
IThis research was supported in part by a grant from the BathSheva de Rothschild Fund for the Advancement of Science and
Technology. 2 Contribution from the Volcani Institute of Agricultural Research, Bet Dagan, Israel, 1969. Series 1588-E. 533
Catechol None Catechol None Catechol None Catechol None
67
20 21 68 10 17 16 21 16 19 18 9 57 24 205 210 230 235
The role of the pH of the external incubating medium was studied first, and a pH of 7.0 was found to give the highest response (Fig. 1). All other experiments were carried out at that pH level. Phenolic compounds, some of which are commonly used as substrates in assays for phenolase activity (3), were dissolved (10-3 M) in 0.1 M phosphate buffer, pH 7.0, and tested for their effect on ethylene production by plugs of green Shamouti orange peel in the system described above. Catechol and 4-methyl catechol both significantly stimulated ethylene produc-
534
FUCHS
FIG. 2. Ethylene induced by catechol. Five plugs of green peel of Shamouti orange fruit were incubated in 2 ml of different concentrations of catechol solutions in 0.1 M phosphate buffer, pH 7.0, in 50-ml sealed flasks. Ethylene was determined after 20 hr of incubation at 20 C, and data are averages of three experiments.
tion by the green peel plugs, whereas p-cresol, guaiacol, p-coumaric acid, resorcinol, and quinol did not affect it (Table I). A 10-2 M concentration of catechol caused the greatest stimulation; concentrations below 10-3 M had no effect on ethylene production; and 10-l M damaged the peel plugs and was inhibitory to the ethylene formation (Fig. 2). Green peel plugs of Clementine (Citrus reticulata B.) responded similarly. Catechol did not affect the ethylene production of avocado (Persea americana) or apple (Pyrus nal/us) fruit plugs. When the color of the Shamouti peel was orange, the ability to produce ethylene and to respond to catechol treatment was less than that with green peel. This is in accordance with the report about the ethylene production rise that was detected with young citrus fruit but not with mature ones (1).
Plant Physiol. Vol. 45, 1970
2, 3-Naphthalenediol (5 X 10-3 M), a specific competitive inhibitor of phenolase (6), completely inhibited the effect of 10-3 M catechol on the ethylene production of green Shamouti peel (Table I). Components of polyphenol oxidases, produced in sweet potato tissue after wounding, were shown to be specific for o-diphenols and could not oxidize monophenols such as p-coumaric acid, p-cresol, and m-cresol (4). From the data reported herein, it is suggested that there may be a phenolase system in the orange peel tissue which is related to the ethylene-forming system. It has already been suggested (10) that phenolase activity may switch on or off the ethylene production in plants. Mapson and Wardale (5) suggested that dihydroxyphenols, such as catechol, are enzymatically oxidized and are thereby competing with the ethylene-forming system for the peroxide generated by the plant tissue. This hypothesis does not hold for the citrus peel system in which catechol accelerates ethylene evolution and an inhibitor of phenolase blocks the effect of catechol. However, in both cases a phenolic compound is a cofactor in the ethylene synthesis system. The data reported here suggest that the role of phenolic substances in the biosynthetic pathway of ethylene in plants needs further clarification. LITERATURE CITED 1. AHARONI, Y. 1968. Respiration of oranges and grapefruits harvested at different stages of development. Plant Physiol. 43: 99-103. 2. GAHAGAN, H. E., R. E. HOLM, AND F. P. ABELES. 1968. Effect of ethylene on peroxidase activity. Physiol. Plant. 21: 1270. 3. HAREL, E., A. M. MAYER, AND Y. SHAIN. 1964. Catechol oxidases from apples, their properties, subcellular location and inhibition. Physiol. Plant. 17: 921-930. 4. HYODO, H. AND I. URITANI. 1967. Properties of polyphenol oxidases produced in sweet potato tissue after wounding. Arch. Biocheimi. Biophys. 122: 299-309. 5. MAPSON, L. W. AND D. A. WARDALE. 1968. Biosyntlhesis of ethylene: enzymes involved in its formation. Biochem. J. 107: 433-442. 6. MAYER, A. M., E. HAREL, AND Y. SHAIN. 1964. 2,3 Naphthalenediol, a specific competitive inhibitor of phenolase. Phytochemistry 3: 447-451. 7. TAKEO T. AND M. LIEBERMAN. 1969. 3-Methylthiopropionaldehyde peroxidase from apples: an ethylene-forming enzyme. Biochim. Biophys. Acta 178: 235-247. 8. VINES, H. M., W. GRIERSON, AND G. J. EDWARDS. 1968. Respiration, internal atmosphere, and ethylene evolution of citrus fiuit. Proc. Amer. Soc. Hort. Sci. 92: 227-234. 9. WILSON, W. C. 1967. Chemical abscission studies of citrus fruit. Proc. Fla. Hort. Soc. 80: 227-231. 10. YANG, S. F. 1967. Biosynthesis of ethylene: ethylene formation from methional by horseradish peroxidase. Arch. Biochem. Biophys. 122: 481-487.
