Spectrophotometric and quantum-chemical study of acid-base and complexing properties of (±)-taxifolin in aqueous solution

Acid-base properties

Neutral form of Tf at pH 1 has one absorption maximum near 285 nm. Spectrum of Tf in strongly acidic solution has a similar profile but it also shows a shoulder in the region of 290–310 nm. A linear relationship between absorbance and concentration for all forms of Tf indicates the absence of the molecular association in solution. Characteristics of the spectral properties of different forms of Tf are given in Table 1. The UV-vis spectra of Tf at different acidities are shown in Figure 1. All raw spectroscopic data are given in the online supplementary material (Tables S1 and S2). The calculations were performed using extinction at high concentration of HCl as extinction of HTf⁺. Determination of the number of main-absorbing species are consistent with the presence of two absorption forms, namely Tf and one tautomer of HTf⁺. The obtained value of equilibrium constant of protonation is 3.14±0.04 and 1.38·10³±110 in logarithmic and absolute units, respectively. The solvation coefficient or m^*-parameter [17] for this process is 3.65±0.08. For comparison, the values of protonation for other flavonoids, such as quercetin and morin are within 9.0–11.0 logarithmic units [18]. The values of solvation coefficient for indoles, amides and tertiary aromatic amines are 1.3, 0.5–0.6 and 1.4, respectively [19].

Figure 1

The UV-vis scanning spectra of Tf obtained at various concentration of HCl and absorbance (308 nm) as a function of log([HCl]), [Taxifolin]=3.16·10⁻⁴m.

Thermodynamic properties of the protonation process were theoretically investigated aiming to check convergence with experimental data. The keto-enol equilibriums of HTf⁺ was assessed through the calculation of absolute and relative energies of each possible tautomer. HTf⁺ has seven tautomeric structures (Scheme 1). The DFT simulations (Table S3) are consistent with the suggestion that N3 (Figure 2A) tautomer is the most energetically favorable structure for HTf⁺. All other isomers are more energetic (≥70 kJ·mol⁻¹) than N3.

Scheme 1

Keto-enol equilibrium of HTf⁺.

Figure 2

Optimization geometry of HTf⁺ (A) and Ni-Tf (B).

Quantum chemical calculations of the protonation constant logK_H were carried out based on the cycle shown in Figure 3 [20]. Using of different computational model with explicit solvation or different solvation model such as COSMO or C-PCM lead to unrealistic theoretical results. In this approximation three main parts of the total free energy in solution ΔΔG^solv. were evaluated: total Gibbs energy in gas ΔG^gas and liquid ΔG^aq. phases and the zero-point energy correction ΔE^zpe.

Figure 3

Thermodynamic cycle for quantum-chemical calculation of logK_H.

Contribution of these parameters to total free energy of reaction is shown in Table 2. As can be seen, the model provides discrepancies between the theoretical and experimental logK_H values of less than 0.2 logarithmic units. Convergence logK_H^calc with logK_H^exp testifies to the correctness of the proposed model of protonation.

Complex formation with Ni(II)

The formation of Ni-Tf complex was indicated by changes in the electronic absorption spectra in solution (Figure 4). The investigation of complexation process in Ni(II)-Tf system was performed under conditions of metal excess. Available pH range for study of this process lie in the range of 7.4–7.8, where Tf exists in mono-anionic form [14]. At pH below 7.4 the interaction Ni^II-Tf is too weak to be measured by a spectrophotometric method, and at pH>7.8 a rapid oxidation of the complex is observed. Interaction between Tf and tris has been not detected under such conditions. Since the ΔA maximum remains invariant at 326 nm (Figure S1) at various nickel concentrations, one might conclude that the complex formation leads to one product (monocomplex species) only with rather negligible contribution from the polynuclear species Ni_nTf_m. Thus, the complexation process of Ni²⁺ with Tf can be described by the equation 1.

Figure 4

The UV-Vis spectra and absorbance at single wavelength (326 nm) for Ni(II)-Tf system; [Taxifolin]=2.24·10⁻⁴m; pH=7.6, I=1 (NaClO₄).

The value of conditional stability constant (logK′) obtained for this system lies within 2.33–2.38 logarithmic units (Table 3). This value remains unchanged within the specified limits for each acidity. This means that H⁺ does not participate in complexation process and a value of n in equation 1 equals 0. The ‘True’ logK cumulative stability constants were obtained from the coefficients of competing reactions [21] (equation 2), in which β_n is the cumulative stability

constant of competing reactions, K′ is conditional stability constant, K is ‘true’ stability constant. For calculation of the coefficient α_m, the equilibrium constant of Ni(II) hydrolysis [22], stability constants of Ni-tris complexes (ML and ML₂ species) [23] and dissociation constant of Tf [14] were used.

The ‘true’ equilibrium stability constant (logK) for Ni-Tf complex is 1.18·10⁵ (or 5.1 logarithmic units). This value characterizes the complex as more stable than other complexes of Tf. For example, in the system Cu(II)-Tf the formed monocomplexes have logK values in the range from −15 to −1 logarithmic units [14].

For estimation of keto-enol equilibrium of Ni-Tf complex, the level Def2-SVP/DFT/PBE0/ Stuttgart RSC 1997/SMD was used. According to [6], Tf can exhibit four possible chelating sites with Ni^II ion (Scheme 2). The DFT calculations the DFT calculations showed that C1 is the most stable tautomer (Figures S2 and 2B) for the Ni^II-Tf structure.

Scheme 2

Keto-enol equilibrium for Ni-Tf complex.

UV-Vis measurements

The Cox-Yates method [25] based on the excess acidity function χ [26] was used to determine the protonation constant (K_H) in strongly acidic solutions (equation 3),

where Ai, AH2L (εH2L), AH3L+ (εH3L+), and AHL− (εHL−) are the absorbances and molar extinction coefficients of the process solution, the free Tf, and its conjugate acid or base, respectively. The number of absorbing species N contributing to the absorbance matrix was estimated with the factor indication function (IND) [27].

Calculations of conditional stability (K′) were performed by nonlinear LSR analysis using the absorbance matrix as raw data [28]. The optimal values for K′ and ε^λ were found from the least squares analysis (equations 4 and 5),

where,

Ab initio study

Ab initio calculations were carried out using the GAMESS US program package [29] with a supercomputer Lomonosov-1 at Moscow State University. The Def2-SVP [30] basis set was applied to H, C and O atoms. Stuttgart RSC 1997 pseudopotentials [31] were applied to Ni(II) for calculation of complexes species. The solvent effects were evaluated using the SMD solvation model [32]. Geometry optimization was performed by density functional theory (DFT). The acid-base equilibrium constants have been calculated using the equations 6–11.

where

Here, RTln([H₂O]) is a free energy change associated with moving a solvent from a standard-state solution phase concentration of 1 m to a standard state of the pure liquid, 55.34 m [33], [34]. E^zpe is calculated harmonic vibrational frequencies to estimate the zero-point energy correction. Free energies for ionization and solvation processes were used for calculations of ΔG reactions both in the solid state and in solution.