Kinetic Study on the Reactivity of Azanone (HNO) toward Cyclic C-Nucleophiles

1. Equipment

UV-Vis absorption spectra were collected using an Agilent 8453 spectrophotometer equipped with a photodiode array detector and thermostated cell holder.

2. Chemicals

The Angeli’s salt stock solution was prepared in 1 mM NaOH. Its concentration was determined by measuring the absorbance at 248 nm (ε = 8.3 · 103 M−1cm−1). The solution was kept on ice. 1-(4-Methoxybenzyl)-2,4-piperidinedione, 2-acetyl-1,3-cyclohexanedione were purchased from Angene Chemical. 1,3-Cyclopentanedione, 2-methyl-1,3-cyclopentanedione, 1,3-cyclohexanedione, 2-methyl-1,3-cyclohexanedione, 1,3-cycloheptanedione, and 2,4-piperidinedione were purchased from Fluorochem, United Kingdom. All other chemicals (of the highest purity available) were sourced from Sigma-Aldrich Corp. All solutions were prepared using deionized water (Millipore Milli-Q system).

3. Kinetic Experiments

The HNO flux was determined from the rate of FlBA oxidation in aerated aqueous solution of Angeli’s salt, monitored at 490 nm. The initial concentration of Angeli’s salt was equal to 20 µM. Due to the scavenging of HNO by O2 and other scavengers, the steady-state concentration of azanone is very low. The HNO dimerization was therefore negligible and was not taken into consideration. HNO released from Angeli’s salt reacts either with the HNO scavenger or with the molecular oxygen to form peroxynitrite, which was detected with the use of the FlBA probe (25 µM). Its reaction with ONOO– results in the formation of fluorescein. The formation of fluorescein was monitored spectrophotometrically by following the increase in its characteristic absorbance at 490 nm. The reaction mixtures contained Angeli’s salt (20 µM), the fluorescein-based monoborate probe FlBA (25 µM), phosphate buffer (50 mM, pH 7.4), dtpa (100 µM), and the HNO scavenger (at an appropriate concentration). In addition, each solution contained 5% (vol.) CH3CN. The rate constants were determined with the assumption that the concentration of molecular oxygen was equal to 225 µM. Each rate constant was determined in at least three independent experiments.

The dataset contains ASCII files with UV-Vis spectra of the reaction mixtures recorded for an incubation time of 10 min. (see Artelska, A.; Rola, M.; Rostkowski, M.; Pięta, M.; Pięta, J.; Michalski, R.; Sikora, A.B. Kinetic Study on the Reactivity of Azanone (HNO) toward Cyclic C-Nucleophiles. Int. J. Mol. Sci. (2021), in press.).

4. Computational Details

Quantum mechanical calculations were performed in the Gaussian G09 suite of programs, Revision E01. Stationary points were found by geometry optimization algorithms with tight convergence criteria except for transition state structure in the reaction of HNO with 1-(4-methoxybenzyl)-2,4-piperidinedione, where default criteria had to be used due to lack of computation convergence. To verify the nature of stationary points, as well as to compute Gibbs free energies, respective frequencies were computed. In calculations, the presence of the water environment was described by the Gaussian default continuum solvation model (IEFPCM). Density functional theory (DFT) functional B2PLYP with Grimme’s D3 dispersion correction (B2PLYP-D3) combined with 6-311+(2df,2p) split valence basis set, was used. The theory level was selected based on the fact that double-hybrid DFT functionals perform well in describing chemical system properties as well as reaction energy barriers, especially when London dispersion corrections are employed.

The file AtomicCoordinatesOfStationaryPoints.txt contains geometries of stationary points obtained at the B2PLYP-D3/6-311+(2df,2p) theory level used for Gibbs free energey calculations in computational studies (see Artelska, A.; Rola, M.; Rostkowski, M.; Pięta, M.; Pięta, J.; Michalski, R.; Sikora, A.B. Kinetic Study on the Reactivity of Azanone (HNO) toward Cyclic C-Nucleophiles. Int. J. Mol. Sci. (2021), in press.).