Evaluation of iron-deficiency stress response of maize

Abstract

A nutrient solution system with siliceous ferrihydrite as the Fe source was developed to screen 20 maize cultivars in a growth chamber for their Fe-deficiency stress tolerances. The Fe-supplying power of the siliceous ferrihydrite was compared with a pure ferrihydrite and FeDTPA in a separate experiment. Attempts were made to collect and purify the phytosiderophore from maize root exudates. Iron from siliceous ferrihydrite, pure ferrihydrite and Fe-chelating resins were used to evaluate the Fe solubilizing capacity of chelate. A high-performance liquid chromatography (HPLC) procedure with a reverse-phase gradient elution system using either sodium or ammonium acetate and 60 % acetonitrile as the eluents, with precolumn derivatization by phenylisothiocyanate (PITC) or 6-araminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC), was developed to qualitatively and quantitatively analyze phytosiderophore. Chromatographic procedures were first developed using hydroxymugineic acid mugineic acid (MA) and deoxymugineic acid (DMA) standards. The twenty maize cultivars showed differences in Fe-deficiency chlorosis in hydroponics culture, as indicated by their visual chlorosis scores, SPAD chlorophyll meter readings and dimethyl sulfoxide extractable chlorophyll levels. The siliceous ferrihydrite suspended in the nutrient solution was as effective as the pure ferrihydrite and FeDTPA in supplying Fe to the maize plants. In evaluation of the phytosiderophore assay procedures, both EDTA and DTPA were able to quantitatively solubilize Fe from the siliceous, pure ferrihydrite or a Fe-chelating hexa-carboxyl resin. The presence of Ca2+ in general reduced the amount of Fe dissolved. The HPLC analysis procedure with PITC as the derivatizing agent and a pH 4.0 ammonium acetate buffer system was the most effective in separating the three phytosiderophore standards. Identity of a suspected phytosiderophore could be confirmed by the reduction or absence of the respective peak of a ferrihydrite-treated sample, and by comparing the UV absorption spectrum of the suspected phytosiderophore with those of the phytosiderophore standards. Standard curves developed by plotting the UV absorption peak area versus concentration of phytosiderophore standards could be used to obtain quantitative estimates of phytosiderophore.