Characteristics of Fe‐deficiency‐induced acidification in subterranean clover

Iron-deficiency-induced acidification is one of the important reactions of plant Fe-deficiency-stress response, but the overall understanding of this reaction is limited. The characteristics of Fe-deficiency-induced acidification of subterranean clover (subclover) (Trifolium brachycalycinum Katzn. and Morley cv. Koala) were studied in this paper. Plants were grown hydroponically under - Fe conditions, and Fe-deficiency-induced acidification was determined using pH-stat, back-titration and chemical equilibrium procedures. Fe-deficiency-induced acidification was undetectable during the first day after Fe-deficiency stress initiation, but the maximum acidification rate was attained by the second day, when plants exhibited visual chlorosis symptoms. The acidification rate was relatively constant with increasing Fe-deficiency chlorosis, suggesting that a critical level of Fe deficiency was needed to trigger acidification, but that once the acidification process was initiated, the intensity of acidification was independent of severity of Fe deficiency. Net H + -release (PR) rate determined using a chemical equilibrium method and net acidity release (AR) rate determined using a back-titration method were practically identical, indicating that Fe-deficiency-induced acidification involved almost entirely the release of free H + , not organic acid. In the assay temperature range of 5 to 35°C, PR rate was highest at about 20°C. Net acidity release rate was almost totally inhibited at pH values <4.5 and increased with increasing assay pH up to pH 9. The pH effect occurred within 30 min of incubation initiation, implying that the effect of pH is probably on the activity of H + transport through the plasma membrane, not on the quantity of responsible protein(s). Cations were required in the incubation solution for Fe-deficiency-induced acidification. Divalent cations in the assay solution resulted in a higher AR rate than monovalent cations, and essential cations resulted in a higher AR rate than non-essential cations, indicating that the relative effectiveness of cations is related to the efficiency of their absorption by plant roots. These results are discussed in relation to their practical significance and the mechanisms of Fe-deficiency-induced acidification.