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Rapid Method for Accurate Determination of Colorless Siderophores and Synthetic Chelates

Dep. of Soil and Water Sciences

Y. Hadar

Dep. of Microbiology and Plant Pathology, Faculty of Agriculture, Hebrew Univ. of Jerusalem, P.O. Box 12, Rehovot, Israel

*Corresponding author (chenyona{at}agri.huji.ac.il).

ABSTRACT

A rapid, simple, and accurate method of determining concentrations of Fe-chelating agents in solution was developed. The assay employs Cu-CAS (chrome azurol S) complex as a testing agent and measures the equilibrium concentration of this complex in the presence of other Fe chelators. The Cu-CAS was found to be advantageous in comparison to other compounds due to: (i) its high extinction coefficient (17 000 M^–1 at pH 5.7) when complexed with Cu, and (ii) the rapid rate at which equilibrium was achieved. The color of the Cu-CAS was measured at a wavelength of 582 nm, and the decrease in absorbance when the Cu is lost to other chelates was measured. Using this method, we successfully measured 50- to 100-nmol quantities of the following chelates: ethylenediaminetetraacetic acid disodium salt (EDTA), ethylenediamine-di(o-hydroxyphenylacetic acid) (EDDHA), desferriferrioxamine-B (DFOB), rhizoferrin, and mugineic acid (MA). We found the method to be accurate and useful in determining concentrations of chelates (not absorbing intensely at 582 nm) including phytosiderophores and microbial siderophores. This method is of particular importance for colorless chelates, which are very difficult to determine otherwise.

Received for publication February 7, 1995.

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