Submitted to: Journal of Chromatography A
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/5/2001
Publication Date: 1/1/2002
Citation: WHEAL, M.S., HELLER, L.I., NORVELL, W.A., WELCH, R.M. REVERSED-PHASE LIQUID CHROMATOGRAPHIC DETERMINATION OF PHYTOMETALLOPHORES FROM STRATEGY II FE-UPTAKE SPECIES BY 9-FLUORENYLMETHYL CHLOROFORMATE FLUORESCENCE. JOURNAL OF CHROMATOGRAPHY A. 2002. v. 942. p. 177-183. Interpretive Summary: Over 40% of the world's soils are deficient in various micronutrient metals especially zinc, copper and iron that result in diminished crop productivity. Food crops produced on these soil types also have reduced nutritional value for humans with respect to the essential micronutrient metals. Phytometallophores are non-protein amino acids released by cereal roots to soils when grown under micronutrient metal deficiency stress conditions; some (e.g., nicotianamine) are involved in re-translocation of micronutrient metals to reproductive organs via the phloem sap. Increasing the amounts of phytometallophores released by cereal roots will increase their efficiency to accumulate micronutrient metals. We have developed a simple and sensitive high-pressure liquid chromatographic technique to quantify the amounts of phytometallophores produced by cereal roots or accumulated in plant tissues. This method will allow researchers to screen nfor and identify micronutrient efficient genotypes of cereal crops and to determine the basic mechanisms of iron and zinc efficiency in cereal crops. It can also be applied to research directed at understanding the homeostatic mechanisms controlling phloem transport of micronutrient metals and in delineating the role of phloem transport in increasing the micronutrient metal density of cereal grains. Such research is required to develop micronutrient rich genotypes of major cereal crops including rice, wheat and maize.
Technical Abstract: An HPLC method to quantitate phytometallophores (phytosiderophores) exuded from roots of barley (Hordeum vulgare L.) growing in nutrient solution culture was developed. 9-fluorenylmethyl chloroformate (FMOC) derivatives of phytometallophores were separated on a C18 reverse phase column using a sodium acetate (pH 7.2) and acetonitrile-methanol gradient over 20 minutes followed by fluorescence detection. Minimum detection limits ranged from 17 to 370 pmol depending on the particular phytometallophore. The effectiveness of this method was demonstrated using the response of barley seedlings to Fe-sufficient and Fe-deficient nutrient solution conditions. Phytometallophores collected in root washings of Fe-deficient barley seedlings increased with plant age while phytometallophore release from Fe-adequate roots was negligible.