Experimental Proton Affinities Research Articles

Rational Design of Cyclopentadiene-Based Super- and Hyperacids Based on Aromaticity.

The design and synthesis of neutral organic superacids have been of interest recently due to their vast applications in chemistry and material sciences, for example, olefinic polymerization, isolation of highly reactive short-lived cations, etc. Cyclopentadiene behaves as a mild organic acid, producing a stable conjugate base by gaining aromaticity and conjugation after deprotonation. To stabilize conjugate bases of organic acids to show superacidities and hyperacidities, we considered aromatic phenyl substituents with cyclopentadiene (mono-, di-, and triphenyl-substituted cyclopentadiene and their cyano derivatives). The MP2, DFT (B3LYP, M06-2X), and CBS-QB3 methods were used to calculate the gas-phase proton affinities of the parent cyclopentadiene, and the DFT methods were chosen for the substituted cyclopentadiene as they yield an experimental proton affinity of cyclopentadiene of 253.66 kcal/mol (Expt. 253.6 ± 1.3 kcal/mol). The stable trisubstituted cyclopentadiene derivative shows gas-phase enthalpies of deprotonation (ΔHacid) of 245 and 239 kcal/mol with DFT B3LYP and M06-2X methods, respectively, with values in the range of hyperacidity. Some of the tautomers of cyclopentadiene derivatives show hyperacidity, with proton affinity values of 205-240 kcal/mol. Triphenyl-substituted cyclopentadiene behaves as a moderate acid but transforms into a superacid after replacing the phenyl group with nitrobenzene, which is a stronger acid than H2SO4 (ΔHacid = 298 kcal/mol). The nucleus-independent chemical shift (NICS) shows the extent of conjugation in the derivatives of cyclopentadienyl anions after deprotonation, which affects the stabilities of conjugate bases, i.e., the acidities of the protonated species. The calculated harmonic oscillator model of aromaticity and NICS indices reveal that the stability of conjugate bases as well as their acidities increase with increasing aromaticity of cyclopentadiene rings after deprotonation in all molecules.

Acid/base properties of α-methyl and gem-dimethyl derivatives of cysteine and serine from the extended kinetic method

The gas-phase proton affinities (PA) for the non-protein α-methyl cysteine (1), gem-dimethyl cysteine (penicillamine) (2), α-methyl serine (3) and 3-methylthreonine (4) have been determined using the extended kinetic method in an ESI-tandem mass spectrometer. Experimental proton affinities of 923.5 ± 9.8, 925 ± 8.5, 932.1 ± 10.1, and 924.5 ± 7.7 kJ/mol were determined for 1–4, respectively. Gas-phase enthalpies of deprotonation (ΔHacid) for 2, 3 and 4 were also determined experimentally to be 1380 ± 9 and 1379 ± 12 and 1378 ± 12 kJ/mol. Hybrid density functional theory calculations at the B3LYP/6–311++G(d,p)//B3LYP/6-31+G(d) level of theory give predictions for the proton affinities of 1–4 and ΔHacid for 2–4 that are in excellent agreement with the measured values. A computed ΔHacid for 1 of 1389 kJ/mol was also determined. Computed acidities for 2–4 were also determined and the agreement with the experimental acidity for 2 is excellent. The computed acidities for 3 and 4 are somewhat larger than the experimental acidities, but are within the experimental error limits, For 1 and 2, the preferred deprotonation site is the SH group on the side chain rather than the COOH group similar to other cysteine analogs that have previously been studied in our lab.

Antioxidant action of phenols: Revisiting theoretical calculations of their thermodynamics

Abstract Theoretical prediction ability of M06-2X functional was tested for thermodynamics of phenol, 15 para and 15 meta phenol derivatives. Calculations were done for gas phase as well as for polar and nonpolar solvents. Although predicted values might be shifted from the experimental ones in the framework of the employed DFT functional and basis set, the calculated and experimental data sets correlate well together. Very good linearity was found especially for the correlation of experimental and theoretical proton affinities. Hammett type correlations between the environments considered were compared. The phenolic C—O bond length was also tested as an alternative substituent effect descriptor while the type and position of the functional group on the aromatic ring have a direct effect on the phenolic bond.

Electron-induced ionization of undeuterated and deuterated benzoic acid isopropyl esters and nicotinic acid isopropyl esters: Some implications for the mechanism of the McLafferty rearrangement.

Electron ionization mass spectra, ionization, and appearance energies and bond energies (as dissociation energies) are reported for benzoic acid-1-methyl-ethyl ester (BAIPE), benzoic acid-1-deutero-1-methyl-ethyl ester (BAIPED1), benzoic acid-2,2,2-trideutero-1-trideuteromethyl-ethyl ester (BAIPED6) as well as nicotinic acid-1-methyl-ethyl ester (NAIPE), nicotinic acid-1-deutero-1-methyl-ethyl ester (NAIPED1), and nicotinic acid-2,2,2-trideutero-1-trideuteromethyl-ethyl ester (NAIPED6). Ionization energies of 9.39 eV for BAIPE, 9.40 eV for BAIPED1, 9.26 eV for BAIPED6 as well as 9.70 eV for NAIPE, 9.79 eV for NAIPED1, and 9.65 eV for NAIPED6 were determined. A gas-phase formation enthalpy of = (-4.10 ± 0.1) eV for BAIPE is calculated as well as = (-3.35 ± 0.1) eV for NAIPE. Molecular ions show two main fragmentation pathways. The first is a classical McLafferty rearrangement, characterized by the transfer of one γ-hydrogen atom from the isopropyl ester chain leading to the ions of the corresponding acid and neutral propene. The second is the double hydrogen transfer from the ester chain leading to the formation of the protonated acid and a C3H5√ allyl radical. For BAIPE, both hydrogen atoms originate from the methyl groups of the aliphatic chain with a probability of ≥98%, whereas the C-1-hydrogen is transferred with a probability of ≤2%. For NAIPE, both hydrogen atoms originate from the methyl groups of the aliphatic chain with a probability of 90%. Experimental proton affinities of PA = (8.75 ± 0.2) eV for benzoic acid and PA = (8.43 ± 0.2) eV for nicotinic acid are derived. For the protonation of the carbonyl group, B3LYP DFT calculations yielded PA = 8.66 eV for benzoic acid and PA = 8.41 eV for nicotinic acid. The overall fragmentation mechanism is explained with the initial formation of a 1,5-distonic ion by transfer of the first hydrogen. For the transfer of the second hydrogen, an intermediate ion/neutral complex is formulated.

Computational insight into the contribution of para-substituents on the reduction potential, proton affinity, and electronic properties of nitrobenzene compounds.

New insight is provided into the chemistry of 12 para-substituted nitrobenzene compounds, using the high-level computational method G3(MP2) and DFT methods. The results show that the chemical properties of the nitrobenzene molecules, such as reduction potential, ionization energy, proton affinity, pKa, interaction energy of the fragments, hyperpolarizability, exaltation index, band gap, UV electron excitation, and QTAIM properties, are controlled by the strong coupling between the nitro group (NO2) and the nature of the various para-substituents via the benzene ring as their conducting link. As the electron donating tendency of the para-substituent increases in the molecules, the electron cloud around the nitro group also increases, resulting in contraction of the N-C bonds and elongation of the N=O bonds, consequently leading to gradually increasing electron conductivity, polarizability, and ionization energy but lower proton affinity, thereby progressively impeding the reduction potential of the molecules. The experimental reduction potential was reproduced to a high degree of accuracy, with a mean absolute deviation (MAD) of 0.048V, depending on the computational method used and the choice of the free energy circle. Additionally, the experimental electron affinity and proton affinity of the 12 molecules were reproduced to a high degree of accuracy. Graphical abstract The correlation of experimental and computational reduction potential of twelve nitrobenzen compounds.

Flue gas adsorption on periodic mesoporous phenylene-silica: a DFT approach.

Periodic mesoporous organosilicas (PMOs) were suggested as potential adsorbents for CO2/CH4 separation because of their large affinities towards CO2 and low interaction with CH4. Herewith, we present a comprehensive computational study on the binding properties of flue gas species with the pore walls of periodic mesoporous phenylene-silica (Ph-PMO) for understanding the possible impact of other gaseous species in the CO2/CH4 separation. The calculations considered three exchange-correlation functionals (PBE, PBE-D2 and M06-2X) based on the density functional theory and the walls of the periodic mesoporous phenylene-silica were modelled within the cluster model approach. The components of the flue gas considered were the diatomic CO, H2, N2, O2 and NO molecules, the triatomic CO2, H2O, H2S and SO2 species, the tetratomic SO3 and NH3 gases and the pentatomic CH4 molecule. The calculated data demonstrate that the presence of H2O, SO2, NH3, H2S and SO3 is a significant threat to CO2 capture by Ph-PMO and suggest that the Ph-PMO material would present high selectivity for CO2 over CH4, CO, H2 or N2 adsorption. The adsorption behaviour of flue gas components in Ph-PMO can be directly related to the experimental proton affinities, basicities or even the polarizabilities of the gaseous molecules.

Vitamin C: an experimental and theoretical study on the gas-phase structure and ion energetics of protonated ascorbic acid.

In order to investigate the gas-phase mechanisms of the acid catalyzed degradation of ascorbic acid (AA) to furan, we undertook a mass spectrometric (ESI/TQ/MS) and theoretical investigation at the B3LYP/6-31 + G(d,p) level of theory. The gaseous reactant species, the protonated AA, [C6 H8 O6 ]H+ , were generated by electrospray ionization of a 10-3 M H2 O/CH3 OH (1 : 1) AA solution. In order to structurally characterize the gaseous [C6 H8 O6 ]H+ ionic reactants, we estimated the proton affinity and the gas-phase basicity of AA by the extended Cooks's kinetic method and by computational methods at the B3LYP/6-31 + G(d,p) level of theory. As expected, computational results identify the carbonyl oxygen atom (O2) of AA as the preferred protonation site. From the experimental proton affinity of 875.0 ± 12 kJ mol-1 and protonation entropy ΔSp 108.9 ± 2 J mol-1 K-1 , a gas-phase basicity value of AA of 842.5 ± 12 kJ mol-1 at 298 K was obtained, which is in agreement with the value issuing from quantum mechanical computations. Copyright © 2016 John Wiley & Sons, Ltd.

A theoretical study of the gas phase (proton affinity) and aqueous (pKa) basicity of a series of 150 pyrazoles

Experimental proton affinities and pKas covering a range of 208 kJ mol−1and 10.3 pKaunits, respectively, have been analyzed theoretically.

Gas-phase acid-base properties of 1-aminocycloalkane-1-carboxylic acids from the extended kinetic method

The gas-phase proton affinities (PA) and gas-phase acidities (GA) for the non-protein amino acids 1-aminocyclopropane-1-carboxylic acid (1), 1-amino-1-cyclobutane-carboxylic acid (2), cycloleucine (3) and 1-amino-1-cyclohexane (4) have been determined using the extended kinetic method in ESI-tandem mass spectrometers. These non-protein amino acids are found in a variety of foods and can compete with other aliphatic amino acids in a variety of biochemical processes. We find a positive trend in proton affinity with an increasing ring size for the four amino acids. Experimental proton affinities of 896±8.0, 913±8.0, 931±8.0, and 933±8.0 were determined for 1–4, respectively. Hybrid density functional theory calculations at the B3LYP/6-311++G(d,p)//B3LYP/6-31+G(d) level of theory give predictions for the proton affinities of 1–4 that are in excellent agreement with the measured values and support the positive trend. In contrast, we find that the gas-phase acidities (ΔHacid) for 1–4 are the same within error. Acidities of 1425±8kJ/mol, 1424±8kJ/mol, 1428±10kJ/mol, and 1423±8kJ/mol were determined for 1–4. Theoretical predictions for the acidities for 1–4 are in excellent agreement with the measured acidities and support the conclusion that there is no trend with changing ring size.

Amine Superbases Stabilized by Extended Hydrogen Bond Networks

Extended hydrogen-bonding networks as a mechanism for creating superbases is explored through six different amine scaffolds: linear acenes, cyclohexane, decalin, triptycene, adamantane, and [2.2]paracyclophane. The gas-phase proton affinities of 21 different potential superbases were computed at the ωB97X-D/6-311+G(2d,p) level. This method was benchmarked against the experimental proton affinities of 44 nitrogen bases. Extended hydrogen-bonding networks, including second- and third-layer hydrogen bonding, led to bases with proton affinities 20 kcal mol(-1) greater than that of bis(dimethylamino)naphthalene. The strongest bases are the decalin base 25 and the adamantane base 31.

Proton affinity of several basic non-standard amino acids

The structures and absolute proton affinities of several arginine (2-amino-3-guanidinopropionic acid, 2-amino-4-guanidinobutyric acid, homoarginine, citrulline and canavanine), histidine (1-methylhistidine and 3-methylhistidine) and lysine (2,3-diaminopropanoic acid, 2,4-diaminobutanoic acid, ornithine, 5-hydroxylysine, canaline and thialysine) homologues and analogues have been estimated using composite G3MP2B3 computational protocol. For a majority of here studied non-standard amino acids the gas-phase proton affinities were established for the first time, while for the others obtained values are used to improve the accuracy of the computational and experimental proton affinities reported previously. In addition, structures and proton affinities are discussed in order to rationalize their biological activity.

Gas‐phase chemistry of ionized and protonated GeF4: a joint experimental and theoretical study

The gas-phase ion chemistry of GeF(4) and of its mixtures with water, ammonia and hydrocarbons was investigated by ion trap mass spectrometry (ITMS) and ab initio calculations. Under ITMS conditions, the only fragment detected from ionized GeF(4) is GeF(3)(+). This cation is a strong Lewis acid, able to react with H(2)O, NH(3) and the unsaturated C(2)H(2), C(2)H(4) and C(6)H(6) by addition-HF elimination reactions to form F(2)Ge(XH)(+), FGe(XH)(2)(+), Ge(XH)(3)(+) (X = OH or NH(2)), F(2)GeC(2)H(+), F(2)GeC(2)H(3)(+) and F(2)GeC(6)H(5)(+). The structure, stability and thermochemistry of these products and the mechanistic aspects of the exemplary reactions of GeF(3)(+) with H(2)O, NH(3) and C(6)H(6) were investigated by MP2 and coupled cluster calculations. The experimental proton affinity (PA) and gas basicity (GB) of GeF(4) were estimated as 121.5 ± 6.0 and 117.1 ± 6.0 kcal mol(-1), respectively, and GeF(4)H(+) was theoretically characterized as an ion-dipole complex between GeF(3)(+) and HF. Consistently, it reacts with simple inorganic and organic molecules to form GeF(3)(+)-L complexes (L = H(2)O, NH(3), C(2)H(2), C(2)H(4), C(6)H(6), CO(2), SO(2) and GeF(4)). The theoretical investigation of the stability of these ions with respect to GeF(3)(+) and L disclosed nearly linear correlations between their dissociation enthalpies and free energies and the PA and GB of L. Comparing the behavior of GeF(3)(+) with the previously investigated CF(3)(+) and SiF(3)(+) revealed a periodically reversed order of reactivity CF(3)(+) < GeF(3)(+) < SiF(3)(+). This parallels the order of the Lewis acidities of the three cations.

Acid–base thermochemistry of gaseous oxygen and sulfur substituted amino acids (Ser, Thr, Cys, Met)

Acid-base thermochemistry of isolated amino acids containing oxygen or sulfur in their side chain (serine, threonine, cysteine and methionine) have been examined by quantum chemical computations. Density functional theory (DFT) was used, with B3LYP, B97-D and M06-2X functionals using the 6-31+G(d,p) basis set for geometry optimizations and the larger 6-311++G(3df,2p) basis set for energy computations. Composite methods CBS-QB3, G3B3, G4MP2 and G4 were applied to large sets of neutral, protonated and deprotonated conformers. Conformational analysis of these species, based on chemical approach and AMOEBA force field calculations, has been used to identify the lowest energy conformers and to estimate the population of conformers expected to be present at thermal equilibrium at 298 K. It is observed that G4, G4MP2, G3B3, CBS-QB3 composite methods and M06-2X DFT lead to similar conformer energies. Thermochemical parameters have been computed using either the most stable conformers or equilibrium populations of conformers. Comparison of experimental and theoretical proton affinities and Δ(acid)H shows that the G4 method provides the better agreement with deviations of less than 1.5 kJ mol(-1). From this point of view, a set of evaluated thermochemical quantities for serine, threonine, cysteine and methionine may be proposed: PA = 912, 919, 903, 938; GB = 878, 886, 870, 899; Δ(acid)H = 1393, 1391, 1396, 1411; Δ(acid)G = 1363, 1362, 1367, 1382 kJ mol(-1). This study also confirms that a non-negligible ΔpS° is associated with protonation of methionine and that the most acidic hydrogen of cysteine in the gas phase is that of the SH group. In several instances new conformers were identified thus suggesting a re-examination of several IRMPD spectra.

Calculation of the enthalpies of formation and proton affinities of some isoquinoline derivatives

Ab initio molecular orbital theory has been used to calculate enthalpies of formation of isoquinoline, 1-hydroxyisoquinoline, 5-hydroxyisoquinoline, and 1,5-dihydroxyisoquinoline as well as some pyridine and quinoline derivatives. The proton affinities of the four isoquinoline derivatives were also obtained. The high-level composite methods G3(MP2), G3(MP2)//B3LYP, G3//B3LYP, and CBS-QB3 have been used for this study, and the results have been compared with available experimental values. For six of the eight studied compounds, the theoretical enthalpies of formation were very close to the experimental values (to within 4.3 kJ · mol −1); where comparison was possible, the theoretical and experimental proton affinities were also in excellent agreement with one another. However, there is an extraordinary discrepancy between theory and experiment for the enthalpies of formation of 1-hydroxyisoquinoline and 1,5-dihydroxyisoquinoline, suggesting that the experimental values for these two compounds should perhaps be re-examined. We also show that popular low cost computational methods such as B3LYP and MP2 show very large deviations from the benchmark values.

Quantum-Chemical Calculations of the Affinity of Neutral Molecules for Protons

Quantum-chemical calculations (B3LYP/6-31++G(d,p)) of proton affinities were performed for molecules of various structures in the gas phase. The errors in calculation results were estimated. The data on the proton affinity of several molecules for which experimental proton affinity values are unavailable were obtained. The hypothesis according to which the structure of heteroassociates in the HF-aprotic organic solvent systems depended on the proton affinity of solvent molecules was substantiated.

Gas-Phase Reactivity of 2,7-Dimethyl-[1,2,4]-triazepine Thio Derivatives toward Cu+Cation: A DFT Study

The gas-phase interactions of 2,7-dimethyl-[1,2,4]-triazepine and its thio derivatives with Cu+ were studied through the use of high-level density functional theory (DFT) calculations. The structure of all possible tautomers and their conformers was optimized at the B3LYP/6-31G(d) level of theory. Final energies were obtained at the B3LYP/6-311+G(2df,2p) level. It has been found that the direct association of Cu+ occurs at the oxygen atom attached to position 3 in the case of the dioxo derivative and at the sulfur atom in all other cases. For the dithio derivatives, the global minimum of the PES corresponds to the structure in which the metal ion bridges between the heteroatom at position 3 and the nitrogen atom at position 4 of the corresponding enolic tautomer, forming a four-membered ring structure; for the dioxo derivative, this conformer competes with the ketone tautomer. Moreover, the isomerization processes leading from the most stable adduct to the other stable conformers were investigated. Among all the considered compounds, the 3,5-dithiotriazepines-Cu+ is found to be the one that associates Cu+ more tightly in the gas phase. The calculated Cu+ binding energies show a good correlation with the experimental proton affinities.

Experimental and theoretical proton affinities of methionine, methionine sulfoxide and their N- and C-terminal derivatives

The proton affinities of methionine, methionine sulfoxide and their derivatives (methionine methyl ester, methionine sulfoxide methyl ester, methionine methyl amide, methionine sulfoxide methyl amide, N-acetyl methionine, N-acetyl methionine sulfoxide, N-acetyl methionine methyl ester, N-acetyl methionine sulfoxide methyl ester, N-acetyl methionine methyl amide and N-acetyl methionine sulfoxide methyl amide) were experimentally determined using the kinetic method, in which proton bound dimers formed via electrospray ionization (ESI) were subjected to collision induced dissociation (CID) in a triple quadrupole mass spectrometer. In addition, theoretical calculations carried out at the MP2/6-311 + G(2d,p)//B3LYP/6-31 + G(d,p) level of theory to determine the global minima of the neutral and protonated species of all derivatives studied, were used to predict theoretical proton affinities. The density function theory calculations not only support the experimental proton affinities, but also provide structural insights into the types of hydrogen bonding that stabilize the neutral and protonated methionine or methionine sulfoxide derivatives. Comparison of the proton affinities of the various methionine and methionine sulfoxide derivatives reveals that: (i) oxidation of methionine derivatives to methionine sulfoxide derivatives results in an increase in proton affinity due to higher intrinsic proton affinity and an increase in the ring size formed through charge complexation of the sulfoxide group, which allows more efficient hydrogen bonding compared to the sulfide group; (ii) C-terminal modification by methyl esterification or methyl amidation increases the proton affinity in the order of methyl amide > methyl ester > carboxylic acid due to improved charge stabilization; (iii) N-terminal modification by N-acetylation decreases proton affinity of the derivatives due to lower intrinsic proton affinity of the N-acetyl group as well as due to stabilization of the attached proton by only one functional group (instead of two functional groups in derivatives containing a free amino group); and (iv) a combination of the above methionine modifications is observed to affect the proton affinity in an additive way. While a number of factors might contribute to discrepancies between the theoretical and experimental proton affinities, a key factor may be that the global minimum of the neutral species is further stabilized by strong intramolecular hydrogen bonding, and this particular conformer may not be sampled during dissociation of the proton bound dimer.

Gas phase protonation of α, β and γ-dicarbonyls: Thermochemistry and structures

An experimental and theoretical study of the protonation of representative dicarbonyl compounds M = 2,3-butanedione (biacetyl), 1, 2,4-pentanedione (acetylacetone), 2, 2,5-hexanedione (acetonylacetone), 3, and methyl-acetoacetate, 4 has been carried out. The experimental proton affinities and protonation entropies have been obtained by the extended kinetic method using the orthogonal distance regression (ODR) treatment. Theoretical proton affinities are calculated at the G2MP2 level of theory while protonation entropies were estimated after a detailed treatment of the internal rotations. The data show that protonation of 1 and 2 is associated with negligible protonation entropies while significant negative values are obtained for molecules 3 and 4. Protonation of 2,3-butanedione, 1a, is associated with a tautomerisation inside the proton transfer complex thus leading to protonated 2-hydroxy-butenone, 1bH +. Protonation thermochemistry of 2,4-pentanedione 2 may be simply rationalized by the protonation of its most stable tautomer, the 4-hydroxy-3-pentene-2-one, 2b, to give its most stable protonated form 2bH + stabilized by a strong intramolecular hydrogen bond. Protonation of 2,5-hexanedione 3a most probably produces a cyclic structure stabilized by a covalent bonding, 3aHc +. The structure of neutral methyl acetoacetate 4 sampled during protonation in mass spectrometry experiments appears to be its diketonic form 4a; its protonation leading to an internally hydrogen bonded stabilized structure 4aH +.

Quantum-chemical study of nitrosonium complexes derived from nitrogen-containing heterocycles

The affinities of a series of azines for nitrosonium ion (ANO+) were calculated by quantum-chemical methods, AM1 and ab initio. The ANO+ values were found to increase as the number of nitrogen atoms in the ring decreases and the donor power of substituents increases; ring fusion also increases ANO+. The best agreement between the calculated and experimental ANO+ values was attained with the use of AM1 calculations and ab initio methods with split polarization or diffuse functions. Substituted pyridines showed linear correlations between the calculated ANO+ values and substituent constants σ or σ+ or overall charges on the NO group in the complex. Linear correlations were also found between the calculated ANO+ values of azines and their experimental proton affinities.

Theoretical and Experimental Investigation of the Energetics of Cis−Trans Proline Isomerization in Peptide Models

The energetics of cis-trans proline isomerization in small peptide models have been investigated using the hybrid density functional theory method B3LYP with a 6-31+G* basis set. The molecules studied are models for the phospho-Ser/Thr-Pro substrate for Pin-1, a peptidyl-prolyl isomerase (PPIase) involved in cell division. Pin-1 requires phosphorylation of a Ser or Thr residue adjacent to a Pro residue in the substrate and catalyzes cis-trans isomerization about the proline amide bond. The dihedral angle that would correspond to the reaction coordinate for isomerization of the omega peptide bond was investigated for several small models. Relaxed potential energy scans for this dihedral angle in N-methylacetamide, 1, N,N-dimethylacetamide, 2, acetylpyrrolidine, 3 and acetylproline, 4, were carried out in 20 degrees steps using the B3LYP/6-31+G* level of theory. In addition, similar scans were carried out for 1-4 protonated on the acetylamide carbonyl oxygen. Optimized structures for 1-4 protonated on the amide nitrogen were also obtained at B3LYP/6-31+G*. Relative proton affinities were determined for each site at various angles along the reaction coordinate for isomerization. The relative proton affinities were anchored to experimental gas phase proton affinities, which were taken from the literature for 1 and 2, or determined in an electrospray ionization-quadrupole ion trap instrument using the extended kinetic method for 3 and 4. Proton affinities of 925 +/- 10 and 911 +/- 12 kJ/mol were determined for 3 and 4, respectively. These studies suggest that the nitrogen atom in these amides becomes the most basic site in the molecule at a dihedral angle of ca. 130 degrees . In addition, the nitrogen atoms in 2-4 are predicted to attain basicities in the range 920-950 kJ/mol, making them basic enough to be the preferred site for hydrogen bonding in the Pin-1 active site, in support of the proposed mechanism for PPIases.