Reaction Pathways of Proton Transfer in Hydrogen-Bonded Phenol–Carboxylate Complexes Explored by Combined UV–Vis and NMR Spectroscopy

Combined low-temperature NMR/UV–vis spectroscopy (UVNMR), where optical and NMR spectra are measured in the NMR spectrometer under the same conditions, has been set up and applied to the study of H-bonded anions A··H··X– (AH = 1-13C-2-chloro-4-nitrophenol, X– = 15 carboxylic acid anions, 5 phenolates, Cl–, Br–, I–, and BF4–). In this series, H is shifted from A to X, modeling the proton-transfer pathway. The 1H and 13C chemical shifts and the H/D isotope effects on the latter provide information about averaged H-bond geometries. At the same time, red shifts of the π–π* UV–vis absorption bands are observed which correlate with the averaged H-bond geometries. However, on the UV–vis time scale, different tautomeric states and solvent configurations are in slow exchange. The combined data sets indicate that the proton transfer starts with a H-bond compression and a displacement of the proton toward the H-bond center, involving single-well configurations A–H···X–. In the strong H-bond regime, coexisting tautomers A··H···X– and A–···H··X are observed by UV. Their geometries and statistical weights change continuously when the basicity of X– is increased. Finally, again a series of single-well structures of the type A–···H–X is observed. Interestingly, the UV–vis absorption bands are broadened inhomogeneously because of a distribution of H-bond geometries arising from different solvent configurations

NMR Study of CHN Hydrogen Bond and Proton Transfer in 1,1-Dinitroethane Complex with 2,4,6-Trimethylpyridine

The intermolecular complex with a CHN hydrogen bond formed by 1,1-dinitroethane (DNE) and 2,4,6-trimethylpyridine (collidine) dissolved in CD₂Cl₂ was studied experimentally by ¹H NMR spectroscopy at 180–300 K. Equilibrium between the molecular CH···N form and the zwitterionic C^–/HN⁺ form was detected in the slow exchange regime in the NMR time scale. No sign of a direct C^–···HN⁺ bond was observed; the ion pair is likely to be held by Coulomb interactions. Moreover, there are indications that the protonated base is involved in the formation of homoconjugated (NHN)⁺ collidine–collidinium hydrogen bonded complexes. The reaction pathway of proton transfer in the DNE–pyiridine complex in a vacuum was studied computationally at the B3LYP/6-31++G­(d,p) level of theory. NMR chemical shifts and coupling constants were calculated for a series of snapshots along the proton transfer coordinate. While the central carbon atom has a pyramidal (sp³) configuration in DNE, it is flat (sp²) in the DNE carbanion. As a result, the most indicative computed NMR parameter reflecting hybridization of a carbon atom appeared to be ¹<i>J</i>_CC, which starts to change rapidly as soon as a structure with a quasi-symmetric C··H··N bond is reached. Couplings within the hydrogen bridge, ¹<i>J</i>_CH, ^1h<i>J</i>_HN, and ²<i>J</i>_CN, can serve as good indicators of the degree of proton transfer

Hydrogen Bonding in Bis(6-amino-1,3-dimethyluracil-5-yl)-methane Derivatives: Dynamic NMR and DFT Evaluation

Three bis­(6-amino-1,3-dimethyluracil-5-yl)-methane derivatives were studied experimentally by variable-temperature ¹H NMR in polar aprotic solutions (CD₂Cl₂, C₅D₅N, C₂D₂Cl₄) and computationally by DFT. The unusual for diarylmethanes coplanar conformation of dimethyluracil rings of each molecule is held by a pair of unequal intramolecular N–H···O hydrogen bonds. We show the presence of two dynamic processes involving breakage/formation of these bonds. First, it is two independent NH₂ group rotations, each coupled to nitrogen inversion. Second, it is uracil ring rotations (ring flips). The thermodynamic parameters (Δ<i>H</i>^‡, Δ<i>S</i>^‡, and Δ<i>G</i>^‡) of both processes were estimated by the full line shape analysis of NMR signals and also by DFT calculations. We demonstrate that, though the ring flips exchange pairs of NH protons, the two processes are not coupled: during the ring flip NH₂ groups do not rotate, and during the NH₂ rotation the rings do not necessarily rotate. Unlike in many other diarylmethanes, the ring flips in the studied compounds are happening stepwise; i.e., the configuration when both rings are “in flight” at the same time is energetically unfavorable (small degree of “cog wheel effect”). The signs of the Δ<i>S</i>^‡ values indicate that the molecular flexibility increases during the NH₂ rotations, but decreases during the ring flips

Solvent and H/D Isotope Effects on the Proton Transfer Pathways in Heteroconjugated Hydrogen-Bonded Phenol-Carboxylic Acid Anions Observed by Combined UV–vis and NMR Spectroscopy

Heteroconjugated hydrogen-bonded anions A···H···X^– of phenols (AH) and carboxylic/inorganic acids (HX) dissolved in CD₂Cl₂ and CDF₃/CDF₂Cl have been studied by combined low-temperature UV–vis and ¹H/¹³C NMR spectroscopy (UVNMR). The systems constitute small molecular models of hydrogen-bonded cofactors in proteins such as the photoactive yellow protein (PYP). Thus, the phenols studied include the PYP cofactor 4-hydroxycinnamic acid methyl thioester, and the more acidic 4-nitrophenol and 2-chloro-4-nitrophenol which mimic electronically excited cofactor states. It is shown that the ¹³C chemical shifts of the phenolic residues of A···H···X^–, referenced to the corresponding values of A···H···A^–, constitute excellent probes for the average proton positions. These shifts correlate with those of the H-bonded protons, as well as with the H/D isotope effects on the ¹³C chemical shifts. A combined analysis of UV–vis and NMR data was employed to elucidate the proton transfer pathways in a qualitative way. Dual absorption bands of the phenolic moiety indicate a double-well situation for the shortest OHO hydrogen bonds studied. Surprisingly, when the solvent polarity is low the carboxylates are protonated whereas the proton shifts toward the phenolic oxygens when the polarity is increased. This finding indicates that because of stronger ion-dipole interactions small anions are stabilized at high solvent polarity and large anions exhibiting delocalized charges at low solvent polarities. It also explains the large acidity difference of phenols and carboxylic acids in water, and the observation that this difference is strongly reduced in the interior of proteins when both partners form mutual hydrogen bonds

Two-Dimensional UV–vis/NMR Correlation Spectroscopy: A Heterospectral Signal Assignment of Hydrogen-Bonded Complexes

We explore in a combined UV–vis/NMR approach the hydrogen-bonded complexes of 2-chloro-4-nitrophenol (CNP) with acetate anion as a model for amino acid side-chain interactions. For the first time, we present two-dimensional UV–vis/NMR correlation spectra, measured simultaneously for the same sample inside of the magnet of a NMR spectrometer. Synchronous and asynchronous 2D plots allow us to monitor the formation of hydrogen-bonded complexes, assign the signals to specific species, and finally estimate the geometry of a hydrogen-bonded 1:1 heteroconjugated anion coexisting with four other anionic species formed in CD₂Cl₂ solution at 180 K. Combined analysis of NMR and UV–vis spectra with the help of previously published hydrogen-bond correlations shows that the hydrogen bond in the heteroconjugate is of the CNP–O^–···HOOCCH₃ type with <i>r</i>(O···O) ≈ 2.48 Å and the average bridging proton displacement from the hydrogen bond center of about 0.18 Å

NMR Studies of the Stability, Protonation States, and Tautomerism of ¹³C- and ¹⁵N-Labeled Aldimines of the Coenzyme Pyridoxal 5′-Phosphate in Water

We have measured the pH-dependent 1H, 13C, and 15N NMR spectra of pyridoxal 5′-phosphate (13C2-PLP) mixed with equal amounts of either doubly 15N-labeled diaminopropane, 15Nα-labeled l-lysine, or 15Nε-labeled l-lysine as model systems for various intermediates of the transimination reaction in PLP-dependent enzymes. At low pH, only the hydrate and aldehyde forms of PLP and the free protonated diamines are present. Above pH 4, the formation of single- and double-headed aldimines (Schiff bases) with the added diamines is observed, and their 13C and 15N NMR parameters have been characterized. For 1:1 mixtures the single-headed aldimines dominate. In a similar way, the NMR parameters of the geminal diamine formed with diaminopropane at high pH are measured. However, no geminal diamine is formed with l-lysine. In contrast to the aldimine formed with the ε-amino group of lysine, the aldimine formed with the α-amino group is unstable at moderately high pH but dominates slightly below pH 10. By analyzing the NMR data, both the mole fractions of the different PLP species and up to 6 different protonation states including their pKa values were obtained. Furthermore, the data show that all Schiff bases are subject to a proton tautomerism along the intramolecular OHN hydrogen bond, where the zwitterionic form is favored before deprotonation occurs at high pH. This observation, as well as the observation that around pH 7 the different PLP species are present in comparable amounts, sheds new light on the mechanism of the transimination reaction

NMR Study of Conformational Exchange and Double-Well Proton Potential in Intramolecular Hydrogen Bonds in Monoanions of Succinic Acid and Derivatives

We present a 1H, 2H, and 13C NMR study of the monoanions of succinic (1), meso- and rac-dimethylsuccinic (2, 3), and methylsuccinic (4) acids (with tetraalkylammonium as the counterion) dissolved in CDF3/CDF2Cl at 300–120 K. In all four monoanions, the carboxylic groups are linked by a short intramolecular OHO hydrogen bond revealed by the bridging-proton chemical shift of about 20 ppm. We show that the flexibility of the carbon skeleton allows for two gauche isomers in monoanions 1, 2, and 4, interconverting through experimental energy barriers of 10–15 kcal/mol (the process itself and the energy barrier are also reproduced in MP2/6-311++G** calculations). In 3, one of the gauche forms is absent because of the steric repulsion of the methyl groups. In all four monoanions, the bridging proton is located in a double-well potential and subject, at least to some extent, to proton tautomerism, for which we estimate the two proton positions to be separated by ca. 0.2 Å. In 1 and 3, the proton potential is symmetric. In 2, slowing the conformational interconversion introduces an asymmetry to the proton potential, an effect that might be strong enough even to synchronize the proton tautomerism with the interconversion of the two gauche forms. In 4, the asymmetry of the proton potential is due to the asymmetric substitution. The intramolecular H-bond is likely to remain intact during the interconversion of the gauche forms in 1, 3, and 4, whereas the situation in 2 is less clear

PO Moiety as an Ambidextrous Hydrogen Bond Acceptor

Hydrogen bond patterns of crystals of phosphinic, phosphonic, and phosphoric acids and their cocrystals with phosphine oxides were studied using ³¹P NMR and single-crystal X-ray diffraction. Two main factors govern these patterns and favor or prevent the formation of cocrystals. The first one is a high proton-accepting ability of the PO moiety in these acids. As a result, this moiety effectively competes with other proton acceptors for hydrogen bonding. For example, this moiety is a stronger proton acceptor than the CO moiety of carboxylic acids. The second factor is the inclination of the PO moiety of both the acids and the oxides to form two hydrogen bonds at once. The peculiarity of these bonds is that they weaken each other to a little degree only. In order to highlight this point, we are using the term “ambidextrous”. These two features should govern the interactions of PO moiety with water and other proton donors and acceptors in molecular clusters, the active sites of enzymes, soft matter, and at surfaces

¹⁵N Nuclear Magnetic Resonance Studies of Acid−Base Properties of Pyridoxal-5‘-Phosphate Aldimines in Aqueous Solution

By use of 15N NMR spectroscopy, we have measured the pKa values of the aldimines 15N-(pyridoxyl-5‘-phosphate-idine)-methylamine (2a), N-(pyridoxyl-5‘-phosphate-15N-idine)-methylamine (2b), and 15N-(pyridoxyl-idine)-methylamine (3). These aldimines model the cofactor pyridoxal-5‘-phosphate (PLP, 1) in a variety of PLP-dependent enzymes. The acid-base properties of the aldimines differ substantially from those of the free cofactor in the aldehyde form 1a or in the hydrated form 1b, which were also investigated using 15N NMR for comparison. All compounds contain three protonation sites, the pyridine ring, the phenol group, and the side chain phosphate (1, 2) or hydroxyl group (3). In agreement with the literature, 1a exhibits one of several pKas at 2.9 and 1b at 4.2. The 15N chemical shifts indicate that the corresponding deprotonation occurs partially in the pyridine and partially in the phenolic site, which compete for the remaining proton. The equilibrium constant of this ring-phenolate tautomerism was measured to be 0.40 for 1a and 0.06 for 1b. The tautomerism is essentially unaltered above pH 6.1, where the phosphate group is deprotonated to the dianion. This means that the pyridine ring is more basic than the phenolate group. Pyridine nitrogen deprotonation occurs at 8.2 for 1a and at 8.7 for 1b. By contrast, above pH 4 the phosphate site of 2 is deprotonated, while the pyridine ring pKa is 5.8. The Schiff base nitrogen does not deprotonate below pH 11.4. When the phosphate group is removed, the pKa of the Schiff base nitrogen decreases to 10.5. The phenol site cannot compete for the proton of the Schiff base nitrogen and is present in the entire pH range as a phenolate, preferentially hydrogen bonded to the solvent. The intrinsic 15N chemical shifts provide information about the hydrogen bond structures of the protonated and unprotonated species involved. Evidence is presented that the intramolecular OHN hydrogen bond of PLP aldimines is broken in aqueous solution. The coupling between the inter- and intramolecular OHN hydrogen bonds is also lost in this environment. The pyridine ring of the PLP aldimines is not protonated in aqueous solution near neutral pH. The basicity of the aldimine nitrogens would be even lower without the doubly negatively charged phosphate group. Protonation of both the Schiff base and pyridine nitrogens has been discussed as a prerequisite for catalytic activity, and the implications of the present findings for PLP catalysis are discussed