Concentrations of Gibberellin in Chilled Hazel Seeds

Abstract

Frankland and Wareing¹ reported that chilling treatments which break the dormancy of hazel (Corylus avellana L.) seeds were also accompanied by an increase in the gibberellin content of the seeds, but that the resultant concentrations of gibberellin appeared to be much too low to account for the subsequent germination of the chilled seeds. On the basis of germination experiments in which hazel seeds were treated with chlorocholine chloride, either during or after a chilling treatment, one of us concluded that gibberellin synthesis in hazel seeds may be an essential part of dormancy-breaking, but that it occurs chiefly after the end of the chilling process². We have therefore re-investigated the gibberellin content of hazel seeds after subjecting them to 28 days' chilling at 5° C in moist sand, and we have made determinations on similarly chilled seeds after further treatment for 2 or 7 days at 20° C. Before extraction, the testa and pericarp of the seeds were discarded and the embryonic axes were removed from the cotyledons for separate extraction. Treatment of the seeds, extraction with 80 per cent methanol and the preparation of the acidic diethyl ether extracts were carried out as described previously². The acidic diethyl ether extracts were fractionated by thin-layer chromatography on ‘Kieselgel G’ using a solvent consisting of ethyl acetate, chloroform and acetic acid (15: 5: 1)³. Strips of material from the thin-layer plates were scraped into tubes, to each of which were added 4 ml. of water, 0.1 ml. of streptomycin (250 U) and ten excised etiolated 5 day old pea epicotyls. Gibberellin was bioassayed by the growth of the pea epicotyl above the first node during 3 days at 22.5° C under continuous illumination of 2,500 lux⁴. A blank thin-layer plate was chromatographed and assayed with each set of bioassays so that an accurate blank value and the least significant differences (P = 0.05) could be calculated⁵. Responses of the bioassay to a British Drug Houses gibberellin preparation (largely gibberellin A₃) which was chromatographed on thin-layer plates indicated that only 10 per cent of the gibberellin was re-extracted from the ‘Kieselgel’ so as to influence the bioassay. Standards of 0.01 and 0.1 nmoles of gibberellin were used, which, after chromatography, gave typical mean stimulations of the upper internodes of the pea epicotyls of 3.9 and 6.9 mm respectively.