Barley Endosperm Bioassay Research Articles

Free and Bound Abscisic Acid and Free Gibberellin-like Substances in Pecan Kernel Tissues during Seed Development

Abstract Pecan [Carya illinoinensis) (Wangenh.) K. Koch] kernel development was characterized by an initial rapid localized expansion of testa and endosperm, which was closely associated with low levels of free and bound abscisic acid (ABA) and with high levels of gibberllin-like (GL) substances. Rapid cotyledon growth began with the termination of testa and endosperm expansion, which was subsequent to a sharp increase in both free and bound ABA. The rate of change for growth in kernel dry weight was highly correlated with the rate of change in levels of both free ABA (R2 = 0.86) and bound ABA (R2 = 0.88). Levels of GL substances (ng/g kernel), as measured by the dwarf pea and cucumber bioassays, were relatively low after the rapid accumulation of kernel dry weight, however, GL substances detected by the barley endosperm bioassay were high during the last 30 days of kernel development. Abscisic acid and GL substances seem to exercise a significant role in seed development.

Some physiological effects of podolactone‐type inhibitors

Podolactone‐type inhibitors, including harringtonolide and lycoricidinol, affect several bioassays at 1 to 10 μM. Although these compounds were less active than abscisic acid in inhibiting growth of coleoptiles of wheat embryos, podolactone A significantly decreased the number of mature spikelets of Lolium temulentum at a lower concentration than abscisic acid. Lycoricidinol was more active than podolactone E, the most active of the podolactones, in three bioassays. Inhibition of growth induced by gibberellic acid or zeatin with podolactone‐type compounds was not competitive. Harringtonolide was a strong inhibitor of growth at higher concentrations but did not affect auxin transport, while podolactone E and lycoricidinol inhibited it. Lycoricidinol, podolactone E and harringtonolide did not retard senescence of leaf‐discs. The promotive effects of gibberellic acid in the barley endosperm bioassay were counteracted by podolactone A.

Relations between the concentration of diffusable and extractable gibberellin-like substances and the alternate-bearing behaviour in apple as affected by chemical fruit thinning

The levels of diffusable and extractable gibberellin-like substances (GA s) of apple fruits were investigated in the alternate-bearing ‘King of the Pippins’ and the regularly fruiting ‘Golden Delicious’, using the barley endosperm bioassay. During 1977 and 1978 there were marked differences between the two cultivars in both the chronological appearance and the absolute amounts of diffusable GA s. The application of fruit-thinning chemicals (carbaryl, ethephon, NAAm) to ‘King of the Pippins’ suppressed the early increase of GA s in diffusates, which normally occurs during the second week after full bloom. The following year, the thinned trees exhibited heavy flower- and fruit-set, while the untreated control trees showed a very poor bloom. The later appearance of the GA peak in diffusates from fruits of ‘Golden Delicious’ (5 weeks after full bloom), was not accompanied by a similar inhibitory effect on subsequent flowering. In contrast to their effect on the levels of diffusable GA s, the thinning-treatments caused no real change in the amount of extractable GA s, which was very similar in fruits from both cultivars.

Effects of Defoliation, Gibberellic Acid and 4-Chlorophenoxyacetic Acid on Growth and Composition of Thompson Seedless Grape Berries

Fifty percent defoliation at berry shatter significantly reduced berry weight of Thompson Seedless grapes. Either gibberellic acid (GA₃) or 4-chlorophenoxyacetic acid (4-CPA) applied to foliated vines produced significantly heavier berries than the control (no defoliation or hormones). GA₃ applied to 50% defoliated vines at berry shatter yielded berries similar in weight to those from control vines, whereas, 4-CPA applied to similar vines had little effect on increasing berry weight. The barley endosperm bioassay of gibberellin (GA)-like substances revealed higher activity in berries from control vines than in berries from defoliated vines sampled one week after berry shatter. GA-like activity in young berries increased in response to exogenous GA₃ application in both control and 50% defoliated vines. No measurable GA-like activity was detected in berry samples collected at véraison. Defoliation reduced the total amount of sugar (total soluble solids) per berry, but not the concentration of total soluble solids. GA₃ and 4-CPA, on the other hand, significantly increased the total sugar per berry, but markedly reduced the level of total soluble solids. The concentration of arginine in mature fruit was significantly less in defoliated vines than control vines, whereas, the levels of malate, tartrate and proline did not differ significantly between these two treatments. GA₃ and 4-CPA significantly increased the level of malate in mature fruits, but reduced the concentrations of proline compared to control fruits. The concentrations of tartrate and arginine in berry juice from vines treated with GA₃ or 4-CPA did not differ significantly from control fruits.

Die Aktivität von einigen freien und konjugierten Gibberellinen im Triticum-Halbkaryopsen- und Amaranthus-Gibberellintest

Summary The biological activity of several gibberellin (GA) conjugates was studied and compared with that of the corresponding free GAs. The following conjugates were included: 3-O- β -D-glucopyranosides of GA 1 , GA 3 and GA 4 ; 13–O- β -D-glucopyranosides of GA 1 , GA 3 and GA 5 ; GA 8 -2-O- β -D-glucopyranoside; 13-O- β -D-glucopyranosyl- GA 5 - β -D-glucopyranosyl ester; GA 3 - β -D-glucopyranosyl ester and GA 3 - α -D-glucopyranosyl ester; N-GA 3 -oyl-glycine, N-GA 3 -oyl-glycylglycine and N-GA 3 -oyl-proline. The activity was determined in the wheat- α -amylase bioassay, i. e., a special modification of the barley endosperm bioassay, and the Amaranthus GA bioassay using sterile techniques. Activities of the free GAs as well as of the conjugates differ in both bioassays. Many of the GA conjugates were found to possess very low activity. Only the 13-O-glucopyranosyl-GA 5 - β -D-glucopyranosyl ester was as active as GA 5 in Triticum , whereas in Amaranthus GA 5 -13-O-glucoside and GA 4 -3–O-glucoside showed relative activities of 7.5 and 15 %, respectively. The GA 3 - β -D-glucopyranosyl ester reached relative activities of 10–15% in both bioassays. According to the present knowledge the activity of GA conjugates as measured in the bioassays is based upon the presence of enzymes hydrolyzing the glucoside, glucosyl ester, and amide-like linkages.

Molecular Structure and Gibberellin Activity. II.* Further Studies on Helminthosporic Acid Analogues

The gibberellin-like activity of twelve synthetic helminthosporins has been examined in the barley endosperm bioassay. (�)-14-Norhelminthospor-7(12)-en-13-oic acid (5) was more potent than GA*9 and elicited similar responses in the assay measuring production of a-amylase, sugar and refractive compounds. Responses to acid (5) and GA*3 were retarded similary by abscisic acid. Log-doseresponse curves after 41 and 65 h incubation were of two types: linear with two C4-cyclohexyl helminthosporins and GA*3; and biphasic with two norhelminthosporins, showing initially a shallow curve then a steep slope at high concentrations. Structure-activity comparisons reveal that an alkyl group at C7 on a helminthosporin bridge contributes significantly to activity, but an olefinic bond has a minor effect. Three different orientations of the bridge carboxyl group did not materially alter activity. Oxygenated cyclohexyl helminthosporins had less biological activity than the deoxygenated analogue, perhaps reflecting a critical orientation of a hydroxyl group in the A ring of the more potent gibberellins.

Hormonal Activity in Detached Lettuce Leaves as Affected by Leaf Water Content

The interrelationship between water deficiency and hormonal makeup in plants was investigated in detached leaves of romaine lettuce (Lactuca sativa L. cv. ;Hazera Yellow'). Water stress was imposed by desiccating the leaves for several hours in light or darkness at different air temperatures and relative humidity. In the course of desiccation, a rise in abscisic acid content and a decline in gibberellin and cytokinin activity were observed by gas-liquid chromatography, by both the barley endosperm bioassay and radioimmunoassay and by the soybean callus bioassay. Gibberellin activity began to decline in the stressed leaves before the rise in abscisic acid, the rate of this decline being positively correlated with the rate of increase in leaf water saturation deficit. Recovery from water stress was effected by immersing the leaf petioles in water while exposing the blades to high relative humidity. This resulted in a decrease in leaf water saturation deficit, a reduction in abscisic acid content, and an increase in gibberellin and cytokinin activity.Application of abscisic acid to the leaves caused partial stomatal closure in turgid lettuce leaves, whereas treatment with gibberellic acid and kinetin of such leaves had no effect on the stomatal aperture. In desiccating leaves, however, gibberellic acid and kinetin treatment considerably retarded stomatal closure, thus enhancing the increase in leaf water saturation deficit. These results suggest that the effect of desiccation in changing leaf hormonal make-up, i.e. a rapid increase in abscisic acid and a decrease in both cytokinin and gibberellin activity, is related to a mechanism designed to curtail water loss under conditions inducing water deficiency.

THE DETERMINATION AND UTILIZATION OF GIBBERELLIC ACID IN GERMINATING BARLEY

A barley endosperm bioassay, utilizing the induction of acid phosphatase in barley endosperm seed halves, has been developed and applied to study both the development of endogenous gibberellin in grain germinating under malting conditions and the utilization of externally applied gibberellic acid. The assay is affected by barley age and variety and these observations may have practical implications. After 24 h germination the level of endogenous gibberellin-like material present in the grain was approximately 1·4 ng/corn. The biological activity of the total gibberellin-like material extracted from grain treated with gibberellic acid fell during germination although significant quantities of applied gibberellic acid had entered the grain during the first 24 h germination.

The Role of Gibberellins in Early Growth and Development of Sugar Beet

Two gibberellin(GA)-like compounds were found in both roots and shoots of sugar beet plants using the barley endosperm bioassay. One had chromatographic properties similar to GA3 and GA1; the other was highly non-polar, relatively inactive in the endosperm assay, and may be a new gibberellin. Presence of the GA3/1-like compound was confirmed with the dwarf rice bio-assay. The quantity of this GA was relatively high in the root compared with the shoot at the 3–4 leaf stage when the first supernumerary cambia are being formed in the root. As plants developed through the 8–9 leaf stage and the 15–16 leaf stage the quantity of GA per unit fresh weight of material decreased. Application of gibberellic acid (GA3) to the roots of young sugar beet plants caused a significant increase in root dry weight shortly after treatment and the rate at which supernumerary cambia were produced was increased. Application of GA3 to a single petiole caused a significant increase in both root and shoot dry weight. GA3 applied to either root or shoot caused a reduction in the rate of leaf formation although total leaf area per plant and shoot dry weight were unaffected. The probable role of GA-like substances in controlling the growth and development of young sugar beet plants is discussed.

Gibberellin-Like Activity of Helminthosporic Acid Analogues

The activity of some analogues of helminthosporins was tested in three gibberellin bioassay systems. Three compounds showed considerable activity in the barley endosperm bioassay, five were slightly active and three were inactive. The response curves were frequently steeper than that of gibberellic acid, and sometimes bimodal. Comparisons of the structures of active and inactive compounds indicate that the hydroxymethyl moiety of natural helminthosporins is not necessary for biological activity. The carboxyl group is required for activity and appears to have favoured spatial requirements. Similar structureactivity relationships were suggested for the Tan-ginbozu rice bioassay but those for the Rumex leaf senescence bioassay were different.

Gibberellin‐like Substances in Developing Barley Grain and Their Relation to Dry Weight Increase

AbstractGibberellin‐like activity of two varieties of barley (Hordeum vulgare L.) at different stages of grain development was determined by barley endosperm bioassay (acid phosphatase bioassay).Activity of the acidic ethyl‐acetate fraction (“free” GAs) in both varieties displayed two maxima, a first maximum at the 9th day and a second one 20 to 21 days after pollination.Activity of the n‐butanol fraction (“bound” GAs) first dropped to a minimum level at the 9th day, then increased to reach a maximum 32 days after pollination, and finally decreased again towards maturity. From the 9th day after pollination, a conversion of “free” GAs to “bound” GAs has probably occurred.From the 12th day after pollination, the curves of rate of dry weight increase and of “free” GAs run nearly parallel, but the latter reached its maximum about 2–4 days earlier than the former.The results indicate that gibberellins may participate in the regulation of the accumulation processes into the barley grain.

THE TIME‐COURSE OF POLAR MOVEMENT OF GIBBERELLIN THROUGH ZEA ROOTS

The polarity of movement of gibberellin through sections cut from near the root tips of Zea mays L. was studied, using methods like those we previously used in roots for auxin and in petioles for auxins, cytokinins, and gibberellic acid (GA‐3). One μg GA‐3 was added in a donor agar block and gibberellin activity in the receiver agar at the opposite end of the section was measured directly with a modified barley endosperm bioassay. The movement of gibberellin was away from the root tip (basipetal) and thus opposite in direction to the polarity of auxin through such root sections. The time‐course of basipetal movement was dissimilar to that for gibberellin or auxin movement through petiole sections. It took 14‐18 hr for gibberellin activity equivalent to 6 ng GA‐3 to collect in the basal receivers on roots. Apical receivers showed activity equivalent to 1.6 ng GA‐3 at 14‐18 hr. Less than 0.01 ng equivalent GA‐3 was collected from sections to which GA‐3 was not added, so the 6 and 1.6 ng were almost entirely due to the added GA‐3. These general conclusions were confirmed with an experiment using 14C‐GA‐3. A decline in activity in receivers was found in some experiments at 18 hr, paralleling earlier results with GA‐3, IAA, and adenine in petioles and IAA in roots.

The Time-Course of Polar Movement of Gibberellin through Zea Roots

The polarity of movement of gibberellin through sections cut from near the root tips of Zea mays L. was studied, using methods like those we previously used in roots for auxin and in petioles for auxins, cytokinins, and gibberellic acid (GA-3). One μg GA-3 was added in a donor agar block and gibberellin activity in the receiver agar at the opposite end of the section was measured directly with a modified barley endosperm bioassay. The movement of gibberellin was away from the root tip (basipetal) and thus opposite in direction to the polarity of auxin through such root sections. The time-course of basipetal movement was dissimilar to that for gibberellin or auxin movement through petiole sections. It took 14-18 hr for gibberellin activity equivalent to 6 ng GA-3 to collect in the basal receivers on roots. Apical receivers showed activity equivalent to 1.6 ng GA-3 at 14-18 hr. Less than 0.01 ng equivalent GA-3 was collected from sections to which GA-3 was not added, so the 6 and 1.6 ng were almost entirely due to the added GA-3. These general conclusions were confirmed with an experiment using 14C-GA-3. A decline in activity in receivers was found in some experiments at 18 hr, paralleling earlier results with GA-3, IAA, and adenine in petioles and IAA in roots.

The Role of Hormones in Controlling the Mechanically Induced Dormancy of Suaeda spp.

AbstractSeeds of several taxa in the genus Suaeda with a mechanically resistant testa were stimulated to germinate with applications: of 1 × 10−4M gibberellic acid (GAg). The barley endosperm bioassay indicated that the non‐dormant taxa, S. macrocarpa, had high initial gibberellin‐like activity while the dormant varieties, S. flexilis and S. vulgaris, had not. Soaking for 2 days increased the gibberellin‐like activity of the dormant taxa. At 20 days hormonal activity was repressed, and few seeds germinated. The barley endosperm bioassay also revealed an inhibitor activity in the dormant varieties. The wheat coleoptile bioassay and UV irradiation of thin layer chromatograms indicated that an inhibitor occurred at the Rf of cochromatographed ABA and that it had similar inhibitory characteristics.

Gibberellin‐like Activity in the Developing Avocado Fruit

AbstractGibberellin‐like activity in avocado (Persea americana) fruit extracts was measured by the barley endosperm bioassay. Fruit tissues were anlyzed separately during fruit growth. The level of the activity found was very high in the endosperm and in the seed coats, but decreased in the latter during fruit growth. No measurable gibberellin‐like activity was detected in the mesocarp or in the embryo. It is assumed that the seed coats are a site of production of gibberellin‐like substances in the avocado fruit.

GA 32 : A Polar Gibberellin with High Biological Potency

A new gibberellin, GA(32), purified from Prunus seed, has three interesting properties: (i) it is the most polar of the 34 gibberellins presently known, (ii) it is the most potent of all gibberellins tested in the barley endosperm bioassay, and (iii) it is very active in promoting the growth of unpollinated apricot ovaries with a normal ratio of length to diameter.

Effect of Stratification and Gibberellin on Seed Germination in Ginkgo biloba

WEST, W. CHARLES, FRANCESCA J. FRATTARELLI, ancd KAREN J. RusSIN (The City College, New York, N.Y. 10031). Effect of stratification ancd gibberellin on seed germinationi in Ginkgo biloba. Bull. Toirey Bot. Club 97: 380-384. 1970.-The effects of stratification ancd exogenous gibberellic acid on seed germination in Ginkgo biloba were followed for a 12-week period. Most nonstratified seeds either remain dormant or decompose during this period, while nearly 100% of stratified seeds germinate within 7-10 weeks. Reilloval of the hard portion of the seed coat produces oiily a slight enhancement in germination of noiistratified seeds. Exogenous application of gibberellic acid to nonstratified seeds enhaniees ger mination to a level that approaches that of stratified seeds. Barley endosperm bioassay data of ethyl acetate extractable gibberellins froim Ginkgo embryos indicate that stratified embryos contain approximately 100 times more GA, equivalents thani nionstratified embryos. It is coneluded that stratificationi anid gibberellin metabolism are related in the germination of Ginkgo seeds, anid that exogenious GA3 will largely substitute for stratification in lionstratified seeds to promote germ-iinationi.

Influence of (2-Chloroethyl) Trimethylammonium Chloride (Cycocel) and Daylength on Gibberellin-like Activity in the Root Exudate of the Poinsettia Euphorbia pulcherrima, Willd. cv. ‘Paul Mikkelsen’1

Abstract Three substances with gibberellin-like activity in the dock and barley endosperm bioassay systems were detected in the root exudate of poinsettia cultivar ‘Paul Mikkelsen’. In the isopropyl ether:acetic acid (95:5, v/v) solvent system on 250 µ layers of silica gel G, these substances had Rf values of 0–0.1, 0.4–0.6, and 0.7–0.8. The extracted exudate material at Rf 0–0.1 generally showed greater activity in plants grown under long day conditions while the relative proportion of the other 2 fractions to the first increased under short day conditions. Following a drench with (2-chloroethyl) trimethylammonium chloride (Cycocel), the activity of all 3 fractions was reduced in comparison to the control. The Rf 0–0.1 material evidenced lower activity in relation to the other 2 fractions in exudate from Cycocel spray-treated plants.

INDUCTION OF FERN SPORE GERMINATION

Light-induced germination of Anemia spores can be inhibited by AMO-1618, a selective inhibitor of gibberellin biosynthesis. The inhibitor has no effect on gibberellin-induced dark germination and its inhibition of light-induced germination can be reversed by supplying gibberellin. Barley-endosperm bioassay of concentrates of medium in which spores are imbibed in light reveals the presence of substances with gibberellin-like activity; assay of medium from dark-imbibed spores does not. Simultaneous exposure of spores to sub-optimal levels of light and gibberellin leads to additivity of effect on germination level. Uptake of labeled gibberellin by spores in light is similar to that in darkness. The implications of these findings for the light-dependent synthesis of a gibberellin-like germination substance are discussed. The bearing of the observations upon understanding the interaction of light and gibberellins in seeds of higher plants is considered.

Native Growth Inhibitors from Citrus Shoots. Partition, Bioassay, and Characterization

A partition method has been devised to separate auxins, gibberellins, and their respective inhibitors in plant extracts. Inhibitors counteracting gibberellin activity have been detected by a modified barley endosperm bioassay. An inhibitor found in young citrus shoots counteracts both auxin and gibberellin activities and behaves during partition and ohromatography like abscisin II.