Kinetic and Mechanistic Studies on the Reaction of the Vitamin B₁₂ Complex Aquacobalamin with the HNO Donor Angeli’s Salt: Angeli’s Salt and HNO React with Aquacobalamin

Abstract

We report the first studies on the reaction between an HNO donor compound and vitamin B12 complexes. Kinetic and mechanistic studies have been carried out on the reaction between the vitamin B12 derivative aquacobalamin (H2OCbl(+)/HOCbl; pKa = 7.8) and the HNO donor Angeli's salt. Studies were carried out with aquacobalamin in excess, since nitrite also reacts with aquacobalamin to form nitrocobalamin (NO2Cbl). At pH <9.90 aquacobalamin reacts directly with the monoprotonated form of Angeli's salt, HN2O3(-), to form nitroxylcobalamin (NO(-)-Cbl(III); NOCbl) and nitrite. At pH >10.80 the reaction instead switches predominantly to a mechanism in which spontaneous decomposition of Angeli's salt to give HNO and nitrite becomes the rate-determining step, followed by the rapid reaction between aquacobalamin and HNO/NO(-) to again give NOCbl. Both reactions proceed with a 1:1 stoichiometry and formation of nitrite is confirmed using the Griess assay.

Studies on the Reaction of Reduced Vitamin B12 Derivatives with the Nitrosyl ­Hydride (HNO) Donor Angeli's Salt: HNO Oxidizes the Transition‐Metal Center of Cob(I)alamin

“…Finally, note that Cbl(II) does not react to any significant extent with cyanide (Figure S6, Supporting Information). The results of these multiple experiments do not support a Cbl(III) intermediate for the reaction of Cbl(II) with AS, despite the fact that Cbl(III) reacts rapidly with AS with a 1:1 stoichiometry to form NOCbl 27. In addition, data simulation using Pro‐Kineticist software also suggest that Cbl(III) is not the intermediate in the reaction between Cbl(II) and AS (data not shown).…”

“…Note that a Cbl(II)/HNO = 1:2 stoichiometry was also found for the nitrite‐free HNO donor Piloty's acid (unpublished results). Given that a substantial fraction of cob(III)alamin [Cbl(III)] exists as aquacobalamin [p K a (H 2 OCbl + ) = 7.834] and that independent experiments have shown that H 2 OCbl + reacts rapidly with AS to give NOCbl,27 this suggests that the Cbl(II) + AS reaction proceeds via a Cbl(III) intermediate. In order to probe this further, experiments were carried out in the presence of an efficient H 2 OCbl + trap cyanide, which reacts rapidly and essentially irreversibly with H 2 OCbl + to form cyanocobalamin [CNCbl; k f = 250 M –1 s –1 and K (CNCbl) ≥ 10 12 M –1 ] 35.…”

“…Several studies report the reactivity of HNO and HNO donors with transition‐metal complexes including porphyrins 19,20. We recently demonstrated that cob(III)alamin (H 2 OCbl + /HOCbl) reacts with AS to form nitroxylcobalamin [nitrosylcobalamin, NO – –Cbl(III)] 27. In this paper we present detailed mechanistic studies on the reaction between reduced cobalamins, cob(II)alamin and cob(I)alamin, with AS.…”

Can nitrocobalamin be reduced by ascorbic acid to nitroxylcobalamin? Some surprising mechanistic findings

“…From the recent literature [ 30 , 31 ] it is known that free · NO can be reduced by ascorbic acid to form HNO, which in turn can react with CblOH 2 to form CblNO. Brasch and coworkers [ 32 ] demonstrated that using Angeli’s salt (HN 2 O 3 − ) as source of HNO, the formation of CblNO can occur at pH > 10.8, where the rate-determining step is the release of HNO by Angeli’s salt, such that no mechanistic details about the mechanism of the reaction between CblOH 2 /CblOH and HNO/NO − could be revealed. At present, it is questionable whether further reduction of · NO to HNO and a direct reaction of CblOH 2 with HNO to form CblNO can account for the results presented in this study.…”

Synthesis and HNO Donating Properties of the Piloty's Acid Analogue Trifluoromethanesulphonylhydroxamic Acid: Evidence for Quantitative Release of HNO at Neutral pH Conditions

“…Given that others have reported that the reaction between HNO and trialkylphosphines is not stoichiometric, the vitamin B 12 derivative hydroxycobalamin (HOCbl⋅HCl) was employed to determine the amount of HNO released. Under alkaline conditions HOCbl (red) reacts stoichiometrically with HNO (1:1) to produce orange nitroxylcobalamin, NOCbl . HOCbl does not react with NO .…”