18th International Symposium on Iron Nutrition and Interaction in Plants
Madrid – May 30-June 3 2016
CHEMICAL FORMS OF CADMIUM, ZINC, AND IRON IN THE PHLOEM SAPS FROM RICE (ORYZA SATIVA
L.) AND CASTOR BEAN (RICINUS COMMUNIS L.)
Tadakatsu Yoneyama * and Tomoko Ariga
Department of Applied Biological Chemistry, the University of Tokyo, Yayoi 1-1-1, Bunkyo-ku, Tokyo 1138657Japan.
* [email protected]
Purpose: During the growth processes of new organs and seed formation in higher plants, all the elements
including essential zinc (Zn) and iron (Fe) in addition to non-essential or toxic cadmium (Cd) are transported
largely via the phloem (Yoneyama et al. 2015). Because of the slight alkalinity (pH 7.5 ‒ 8.5) and high
phosphate concentrations (mM level) of the phloem saps, the cationic metal ions (Cd2+, Zn2+, Fe2+, Fe3+) must
be stabilized by making metal-chelate complexes; otherwise, these ions might be precipitated with phosphate
and deposited on the cell organelles and membranes. We first identified the chemical forms of these metals in
the phloem saps from rice, a graminaceous plant. We also re-examined them in the phloem sap from castor
bean, a non-graminaceous plant.
Collection of phloem saps and speciation analysis of their metals: Phloem sap (1 ‒ 3 μL) was collected
from the leaf-sheaths of mature rice leaves as the droplets from the laser-severed stylets of a planthopper as
first described by Kawabe et al. (1980). The phloem sap (20 ‒ 50 μL) from the castor bean stems near the
petioles of the mature leaves was collected by incision with a razor blade, as previously outlined by Hall and
Baker (1972). The phloem sap samples were separated into 0.25 or 0.5 mL fractions by size-exclusion
chromatography (SEC) with an elution buffer of pH 8.0, and their metal contents were determined using
graphite furnace atomic absorption spectrometry. For some samples, a SEC‒ICP-MS hyphenated system was
adopted. The contents of protein-bound metals were estimated from the differences between samples with
and without pre-treatment by Proteinase K. The chemical forms of Zn and Fe in the SEC fractions were
identified using electrospray ionization time-of-flight mass spectrometry. Metal-chelating compounds,
nicotianamine (NA), 2’-deoxymugineic acid (DMA), phytochelatines (PCs) and citrate, were measured using a
capillary electrophoresis-mass spectrometer.
Possible chemical forms identified: The Cd in phloem saps from the rice and castor bean plants treated
with 1 μM Cd was largely bound to proteins and potentially to SH compounds such as PCs (Kato et al. 2010).
The Zn forms in the phloem saps from both plants were largely in a Zn-NA complex, with some Zn-histidine in
rice (Kato et al. 2010). The Fe forms in the phloem sap from rice plants were a Fe-DMA complex and some
Fe was protein-bound (Nishiyama et al. 2012) and Fe-citrate (Kato et al. 2010); those in the phloem sap from
castor bean plants were largely Fe-NA complexes (Hazama et al. 2015), with some bound to citrate and
proteins (Ariga, unpublished). The concentrations of chelating compounds in rice phloem sap were DMA 152
μM, NA 66 μM, PC2 2.7 μM and citrate (free) 1.0 μM (Kato et al. 2010, Ando et al. 2013), and those in castor
bean phloem sap were NA 180 μM, PC2 3.4 μM and citrate (free) 87 μM (Hazama et al. 2015).
Keywords. Castor bean, Chemical forms, Metals, Phloem sap, Rice.
REFERENCES: Ando et al 2013 Func Plant Biol 40: 89. Hall and Baker 1972 Planta 106: 131. Hazama et al
2015 Physiol Plant 154: 243. Kato et al 2010 Soil Sci Plant Nutr 56: 839. Kawabe et al 1980 Plant Cell Physiol
21: 1319. Nishiyama et al 2012 Plant Cell Physiol 53: 381. Yoneyama et al 2015 Int J Mol Sci 16:19111.
ACKNOWLEDGEMENTs: We thank University of Tokyo phloem-sap metal research group including Mariyo
Kato, Reiko Nishiyama, Yuko Ando, Kenji Hazama, Hiroaki Hayashi, Shinji Nagata, and Shuichi Yanagisawa.
Preferred Presentation format: ORAL.
Selected Sessions: Fe acquisition, transport and distribution in plants.
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