Characterization of the Initial Intermediate Formed during Photoinduced Oxygenation of the Ruthenium(II) Bis(bipyridyl)flavonolate Complex

A ruthenium­(II) flavonolate complex, [Ru^II(bpy)₂fla]­[BF₄], was synthesized to model the reactivity of the flavonol dioxygenases. The treatment of dry CH₃CN solutions of [Ru^II(bpy)₂fla]­[BF₄] with dioxygen under light leads to the oxidative O-heterocyclic ring opening of the coordinated substrate flavonolate, resulting in the formation of [Ru^II(bpy)₂(carboxylate)]­[BF₄] (carboxylate = <i>O</i>-benzoylsalicylate or benzoate) species, as determined by electrospray ionization mass spectrometry. Moderation of the excitation and temperature allowed isolation and characterization of an intermediate, [Ru^II(bpy)₂bpg]­[BF₄] (bpg = 2-benzoyloxyphenylglyoxylate), generated by the 1,2-addition of dioxygen to the central flavonolate ring

Characterization of the Initial Intermediate Formed during Photoinduced Oxygenation of the Ruthenium(II) Bis(bipyridyl)flavonolate Complex

A ruthenium­(II) flavonolate complex, [Ru^II(bpy)₂fla]­[BF₄], was synthesized to model the reactivity of the flavonol dioxygenases. The treatment of dry CH₃CN solutions of [Ru^II(bpy)₂fla]­[BF₄] with dioxygen under light leads to the oxidative O-heterocyclic ring opening of the coordinated substrate flavonolate, resulting in the formation of [Ru^II(bpy)₂(carboxylate)]­[BF₄] (carboxylate = <i>O</i>-benzoylsalicylate or benzoate) species, as determined by electrospray ionization mass spectrometry. Moderation of the excitation and temperature allowed isolation and characterization of an intermediate, [Ru^II(bpy)₂bpg]­[BF₄] (bpg = 2-benzoyloxyphenylglyoxylate), generated by the 1,2-addition of dioxygen to the central flavonolate ring

Roles of AMF inoculation in plant tolerance to Pb.

<p>AMF inoculation increased root Pb concentration but inhibited Pb root-to-leaf translocation. Mycorrhizal plants had higher leaf SOD, APX and GPX activities but lower leaf H₂O₂ and MDA contents compared with non-mycorrhizal plants. AMF inoculation increased Chl contents and gas exchange capacity, and protected the PSII reaction center under Pb stress conditions. AMF symbiosis causes physiological changes in plant aerial parts probably through affecting nutritional status, hormonal balance and/or secondary metabolism, and the translocation of small signaling molecules.</p

The SOD activity (a), POD activity (b), CAT activity (c), APX activity (d), GPX activity (e), and GR activity (f) in <i>R</i>. <i>pseudoacacia</i> leaves were affected by Pb stress levels and AMF inoculation.

<p>NM, non-inoculated control; Fm, inoculated with <i>F</i>. <i>mosseae</i>; and Ri, inoculated with <i>R</i>. <i>intraradices</i>. The results are shown as the mean (n = 6) ± SD. The same letter indicates no significant difference among treatments (Duncan’s test, P < 0.05). Two-way ANOVAs were used to determine the significance of the effects of the Pb level (Pb), AMF inoculation (AMF), and their interactions (Pb × AMF) on those parameters, as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0145726#pone.0145726.s006" target="_blank">S4 Table</a>.</p

Pb concentrations in leaves (a) and roots (b), and the TF (c) of <i>R</i>. <i>pseudoacacia</i> were affected by Pb stress levels and AMF inoculation.

<p>NM, non-inoculated control; Fm, inoculated with <i>F</i>. <i>mosseae</i>; and Ri, inoculated with <i>R</i>. <i>intraradices</i>. The results are the mean (n = 6) ± SD. The same letter indicates no significant difference among treatments (Duncan’s test, P < 0.05). Two-way ANOVAs were used to determine the significance of the effects of Pb stress (Pb), AMF inoculation (AMF), and their interactions (Pb × AMF) on the Pb concentration and TF of black locust seedlings, as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0145726#pone.0145726.s003" target="_blank">S1 Table</a>.</p

Mycorrhizal colonization (MC) of <i>R</i>. <i>pseudoacacia</i> was affected by Pb stress levels.

<p>NM, non-inoculated control; Fm, inoculated with <i>F</i>. <i>mosseae</i>; and Ri, inoculated with <i>R</i>. <i>intraradices</i>. The results are reported as the mean (n = 6) ± SD. The same letter indicates no significant difference among treatments (Duncan’s test, P < 0.05). Two-way ANOVAs were used to determine the significance of the effects of the Pb level (Pb), AMF inoculation (AMF), and their interactions (Pb × AMF) on the MC of black locust seedlings, as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0145726#pone.0145726.s003" target="_blank">S1 Table</a>.</p

Fv/Fm (a), ΦPSII (b), qN (c), qP (d), Pn (e), g_s (f), C_i (g), T_r (h) and WUE (i) in <i>R</i>. <i>pseudoacacia</i> leaves were affected by Pb stress levels and AMF inoculation.

<p>NM, non-inoculated control; Fm, inoculated with <i>F</i>. <i>mosseae</i>; and Ri, inoculated with <i>R</i>. <i>intraradices</i>. The results are represented as the mean (n = 6) ± SD. The same letter indicates no significant difference among treatments (Duncan’s test, P < 0.05). Two-way ANOVAs were used to determine the significance of the effects of Pb stress (Pb), AMF inoculation (AMF), and their interactions (Pb × AMF) on those parameters, as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0145726#pone.0145726.s004" target="_blank">S2</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0145726#pone.0145726.s005" target="_blank">S3</a> Tables.</p

The H₂O₂ content (a) and MDA content (b) in <i>R</i>. <i>pseudoacacia</i> leaves were affected by Pb stress levels and AMF inoculation.

<p>NM, non-inoculated control; Fm, inoculated with <i>F</i>. <i>mosseae</i>; and Ri, inoculated with <i>R</i>. <i>intraradices</i>. The results are the mean (n = 6) ± SD. The same letter indicates no significant difference among treatments (Duncan’s test, P < 0.05). Two-way ANOVAs were used to determine the significance of the effects of the Pb level (Pb), AMF inoculation (AMF), and their interactions (Pb × AMF) on those parameters, as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0145726#pone.0145726.s007" target="_blank">S5 Table</a>.</p

Mechanistic Investigations of Photoinduced Oxygenation of Ru(II) Bis-bipyridyl Flavonolate Complexes

We previously reported that a Ru-bound flavonolate model of flavonol dioxygenases, [Ru^II(bpy)₂(3-hydroxyfla)]­[PF₆], photochemically reacts with dioxygen in two different manners. Broad-band excitation generates mixtures of products characteristic of 1,3-addition of dioxygen across the central pyrone ring, as is observed in enzymatic reactions. However, low temperature excitation at wavelengths longer than 400 nm generates a unique Ru-bound 2-benzoatophenylglyoxylate product resulting from a 1,2-dioxetane intermediate. Herein, we investigate this reactivity in a series of Ru­(II)­bis-bipyridyl flavonolate complexes [Ru^II(bpy)₂(3-hydroxyfla^R)]­[PF₆] (bpy = 2,2′-bipyridine; fla = flavonolate; R = <i>p</i>-OMe (<b>1</b>), <i>p</i>-Me (<b>2</b>), <i>p</i>-H (<b>3</b>), <i>p</i>-Cl (<b>4</b>)), and [Ru^II(bpy)₂(5-hydroxyfla)]­[PF₆] (<b>5</b>). The complexes’ structures, photophysical and electrochemical properties, and photochemical reactivity with oxygen were investigated in detail. Two different reaction product mixtures, from 1,2- and 1,3-additions of dioxygen, are observed by illumination into distinct excitation/emission manifolds. By analogy to previous reports of excited state intramolecular proton transfer, the two manifolds are attributed to tautomeric diradicals that predict the observed reactivity patterns