Abstract

Low-molecular-weight borate complexes were isolated from canola (Brassica napus) and wheat (Triticum aestivum) phloem exudates, as well as the cytoplasm of the fresh-water alga Chara corallina, and identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Phloem exudate was collected from field-grown canola inflorescence stalks by shallow incision, while wheat phloem exudate was collected by aphid stylectomy. Chara cytoplasm was collected by careful manual separation of the cell wall, vacuole, and cytosolic compartments. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry showed the presence of isotopic borate complexes, at mass-to-charge ratio of 690.22/691.22 in the canola and wheat phloem and at 300.11/301.11 in canola phloem and Chara cytoplasm. Using reference compounds, the borate complexes with mass-to-charge ratio 690.22/691.22 was identified as a bis-sucrose (Suc) borate complex in which the 4,6-hydroxyl pairs from the two alpha-glucopyranoside moieties formed an [L(2)B](-1) complex. Further investigation using liquid chromatography electrospray ionization triple quadrupole mass spectrometry analysis confirmed the presence of the bis-Suc borate complex in wheat phloem with a concentration up to 220 microm. The 300.11/301.11 complex was putatively identified as a bis-N-acetyl-serine borate complex but its concentration was below the detection limits of the liquid chromatography electrospray ionization triple quadrupole mass spectrometer so could not be quantified. The presence of borate complexes in the phloem provides a mechanistic explanation for the observed phloem boron mobility in canola and wheat and other species that transport Suc as their primary photoassimilate.

Highlights

  • Low-molecular-weight borate complexes were isolated from canola (Brassica napus) and wheat (Triticum aestivum) phloem exudates, as well as the cytoplasm of the fresh-water alga Chara corallina, and identified using matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry

  • A further investigation was initiated to determine whether other intracellular ligands exist, and to do this, the fresh-water alga Chara corallina was used as it has very large cells enabling the separation of the cell wall, cytoplasm, and vacuole, thereby enabling the collection of pure cytosolic fluid

  • Matrix-assisted laser desorption/ionization time-offlight mass spectrometry (MALDI-TOF-MS) identified two [L2B]21 ions in canola phloem sap with mass-tocharge ratio (m/z) values of 300.11 and 301.11, and 690.22 and 691.22 with a natural abundance of 10B:11B (20%:80%), confirming the existence of borate complexes (Figs. 1 and 2)

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Summary

Introduction

Low-molecular-weight borate complexes were isolated from canola (Brassica napus) and wheat (Triticum aestivum) phloem exudates, as well as the cytoplasm of the fresh-water alga Chara corallina, and identified using matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry. Matrix-assisted laser desorption/ionization time-offlight mass spectrometry showed the presence of isotopic borate complexes, at mass-to-charge ratio of 690.22/691.22 in the canola and wheat phloem and at 300.11/301.11 in canola phloem and Chara cytoplasm. Further investigation using liquid chromatography electrospray ionization triple quadrupole mass spectrometry analysis confirmed the presence of the bis-Suc borate complex in wheat phloem with a concentration up to 220 mM. The presence of borate complexes in the phloem provides a mechanistic explanation for the observed phloem boron mobility in canola and wheat and other species that transport Suc as their primary photoassimilate. Due to its close evolutionary relationship to higher plants (Manhart and Palmer, 1990), the likelihood of similar complexes occurring in higher plants would be high

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