Enhancement of Gibberellin-Induced Phenomena H. M . CATHEY, N . W . STUART, V . K. TOOLE, a n d S A M ASEN
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Crops Research Division, U. S. Department of Agriculture, Beltsville, Md.
Adjusting the solution containing gibberellin t o p H 3 . 2 w i t h c i t r a t e - p h o s p h a t e buffer p r o m o t e d much h i g h e r p e r c e n t a g e g e r m i n a t i o n o f lettuce a n d Lepidium t h a n did unbuffered w a t e r s o l u t i o n . W h e n a d d e d t o l o w concentrations of a p p l i e d gibberellin, hydrangenol (4',8-dihydroxy-3,4-di hydroisocoumarin), i s o l a t e d f r o m t h e l e a v e s of Engel's W h i t e h y d r a n g e a s , , e n h a n c e d t h e stem e x t e n s i o n o b s e r v e d w i t h mutant d w a r f m a i z e , p e a s , b e a n s , chrysanthemums, a n d h y d r a n g e a s . H y d r a n g e n o l e x h i b i t e d little o r n o g r o w t h - r e g u l a tion activity when applied alone or in combina t i o n w i t h h i g h concentrations o f g i b b e r e l l i n . O t h e r chemicals affecting t h e g r o w t h - p r o m o t i n g a c t i v i t y o f g i b b e r e l l i n a r e cinnamic a c i d , p coumaric a c i d , a n d β - h y d r o x y e t h y l h y d r a z i n e .
Jhe rate and amount of growth of certain plants can be modified by applications of aqueous solutions of gibberellic acid. Often high concentrations of gibberellin are necessary for effectiveness, or the limited tissue area for uptake greatly reduces the usefulness of applied gibberellin. This report deals with factors influencing the uptake and activity of gibberellin applied to seeds and plants. Effect of Surfactants on Gibberellin Tween 20 (nonionic, polyoxylethylene sorbitan monolaurate) was reported as the only surfactant among several tested which increased the effectiveness of the gibberellin solution over that of a solution of gibberellin in water alone when applied to dwarf maize (15) and bean (5). No consistent response from adding Tween 20 to the gibberellin solution has been observed by the authors. Tween 20 aided the spraying and pipetting operations, but apparently was not associated with enhancement of chrysanthemum and hydrangea growth. This area of research needs further study. Effect of Chemical Structure of Gibberellins and Derivatives on Vlant Growth The gibberellins were defined by Phinney and West (16) as substances pos sessing the same carbon skeleton as gibberellin A , gibberellic acid, or one very 135 3
GIBBERELLINS Advances in Chemistry; American Chemical Society: Washington, DC, 1961.
ADVANCES I N CHEMISTRY SERIES
Figure 1.
Elongation of second leaf sheath of d-1 maize follouAng application of various gibberellins (0.1 μg.) to base of first leaf sheath
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Left to right. Untreated; gibberellic acid A*; mixture of gibberellins Ai and A»; potassium gibberellate; zinc gibberellate; methyl ester of gibberellic acid; and allogibberic acid closely related to it, and biologically active in stimulating cell division, cell elonga tion, or both in plants. Various derivatives of gibberellic acid have been made to determine if its activity could be enhanced. Salt. When compared on a molar basis, the mineral ion salts (ammonium, calcium, rubidium, copper, silver, lead, manganese, cobalt, potassium, and sodium, and cyclohexylamine salt) were as effective as the free gibberellic acid in promot ing stem elongation (10,14). As shown in Figure 1, the potassium and zinc salts of A were as active as the acid in promoting the growth of d-1 dwarf maize. Acyl. The acetyl, butyryl, benzoyl, and diacetyl derivatives of gibberellin were reported by Moffatt and Radley (14) to be as active as the acid when applied to the roots or leaves of pea. Ester. Esterification of the hydroxyl group of gibberellin A did not reduce biological activity, and none was more active than the acid (14). Straight aliphatic esters prepared by esterifying the carboxyl group were relatively inactive when assayed on seed (10) and growing plants (4). As an example, the methyl ester of gibberellic acid was inactive on cucumber seedlings (10), on d-1 maize (Figure 1), and on the induction of parthenocarpic fruit set of tomato (4), but was slightly active when applied to dormant lettuce seed (17) and to pea seedlings grown in water solutions containing the ester (14). The observed activity may have resulted from the presence of ethanol (19), or from the slow hydrolysis of the ester into the acid. The complete lack of activity shown by acyl esters may have been due to their low solubility. Although the η-butyl ester had little effect on the growth of cucumber seed lings, butyl Cellosolve ester was almost as effective as the acid (10). Since the Cellosolve esters are relatively insoluble in water, hydrolysis may have occurred. Acids. All the previous comparisons were made with gibberellin A . Other gibberellins have been isolated, six from the metabolic products of the fungus Gibberella fujikuroi (Saw) Wr. (Αχ, A , A , A , A , and A ) and four from the immature seed of Phaseolus multiflorus Lem. (Αχ, A , A , and A ) (8, IS).' The biological activity of the first four gibberellins has been characterized (4, 6, 10, 11, 20). On most plant processes, gibberellin A was the most active, whether it was assayed on d-1 mutant maize (Figure 2), Black Valentine snap bean (4), lettuce, or hydrangea (Figure 2). The remaining gibberellins were less effective in the following order: Αχ ^ A > A . Stem extension of cuttings of Fred Shoesmith chrysanthemums treated with the four gibberellins was similarly affected. 3
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136 GIBBERELLINS Advances in Chemistry; American Chemical Society: Washington, DC, 1961.
Gibberellin-lnduced
Phenomena
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CATHEY ET A L .
Figure 2.
Elongation following application of gibberellins A
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1 μg. per plant Top. To d-l maize Middle. 3 times 5 days apart to EngeVs White hydrangeas Bottom. One time to Marketer cucumber Roots on cuttings treated with A were developed as well as or better than those on untreated cuttings. The other gibberellins tended to retard root development. The only exception to the order as measured by stem extension was with cucurbits {10-12, 2 0 ) , where gibberellin A was always the most active, followed by A , A i , and A 4
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137 GIBBERELLINS Advances in Chemistry; American Chemical Society: Washington, DC, 1961.
advances in chemistry
series
Enhancement of Gibberellin Effects in Seed Germination Gibberellin A* was 50 to 100 times more effective than the other gibberellins in promoting the germination of lettuce and tobacco seed (11) in darkness. Although gibberellin A was the most active on the germination of lettuce seed, A was used for extended tests because larger amounts were available. Toole and Cathey (19) compared two seed kinds— Grand Rapids lettuce and Lepidium virginicum L.—in germination tests with gibberellin A . The germina tion of Grand Rapids lettuce increased gradually with increasing concentrations of gibberellin; the optimum concentration was l O ^ M . In contrast, Lepidium re quired a higher and very restricted range of gibberellin concentration (2.5 X l O ^ M ) for promotion of germination. Lower concentrations had no apparent effect, whereas higher concentrations caused the seedlings literally to "pop out" of the seed coat with no development of the radicle. Potassium gibberellate was inactive on Lepidium at all concentrations tested from 10~ to ICHM. Gibberellic acid always bleached the seed coat because of the acidity. The p H of the solution (2.5 X 10" M) of the potassium gibberellate was 6.0, whereas that of the acid was 3.2. When the potassium salt was dissolved in a citrate (0.005M) and phosphate (0.01M) buffer at p H 3.5, it promoted seed germination as well as did the acid. Solutions buffered at a higher p H were ineffective in promoting germination. The buffer alone did not affect the germina tion of Lepidium seed held in darkness but greatly increased the sensitivity of the seed to light. Solutions of varying concentrations of gibberellin weakly buffered at p H 3.2 were much more effective at a lower and over a broader range in promoting the germination of lettuce and Lepidium seeds than were those that had inbibed unbuffered gibberellin solutions (Figure 3). 4
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