Intramolecular Interactions versus Hydration Effects on p

Article pubs.acs.org/JPCA

Intramolecular Interactions versus Hydration Effects on p‑Guanidinoethyl-phenol Structure and pKa Values Najoua Derbel,*,† Igor Clarot,*,‡,§ Maxime Mourer,‡,§ Jean-Bernard Regnouf-de-Vains,‡,§ and Manuel F. Ruiz-López*,‡,§ †

LSAMA, University of Tunis - El Manar, Campus Universitaire, 2092, Tunis, Tunisia Université de Lorraine, SRSMC, UMR 7565, Vandoeuvre-lès-Nancy, F-54506, France § CNRS, SRSMC, UMR 7565, Vandoeuvre-lès-Nancy, F-54506, France ‡

S Supporting Information *

ABSTRACT: We analyze the structure, hydration, and pKa values of p-guanidinoethyl-phenol through a combined experimental and theoretical study. These issues are relevant to understand the mechanism of action of the tetrameric form, the antibacterial compound tetra-p-guanidinoethyl-calix[4]arene (Cx1). The investigated system can also be useful to model other pharmaceutical drugs bearing a guanidine function in the vicinity of an ionizable group and the effect of arginine on the pKa of vicinal ionizable residues (in particular tyrosine) in peptides. The p-guanidinoethyl-phenol monomer (mCx1) has two ionizable groups. One important particularity of this system is that it exhibits high molecular flexibility that potentially leads to enhanced stabilization in folded structures by direct, strong Coulombic interactions between the ionizable groups. The first pKa corresponding to ionization of the −OH group has experimentally been shown to be only slightly different from usual values in substituted phenols. However, because of short-range Coulombic interactions, the role of intramolecular interactions and solvation effects on the acidities of this compound is expected to be important and it has been analyzed here on the basis of theoretical calculations. We use a discretecontinuum solvation model together with quantum-mechanical calculations at the B3LYP level of theory and the extended 6311+G(2df,2p) basis set. Both intra- and intermolecular effects are very large (∼70 kcal/mol) but exhibit an almost perfect compensation, thus explaining that the actual pKa of mCx1 is close to free phenol. The same compensation of environmental effects applies to the second pKa that concerns the guanidinium group. Such a pKa could not be determined experimentally with standard titration techniques and in fact the theoretical study predicts a value of 14.2, that is, one unit above the pKa of the parent ethyl-guanidinium molecule. Scheme 1. Structures of p-Guanidinoethyl-phenol Monomer (mCx1) and of tetra-p-Guanidinoethyl-calix[4]arene (Cx1), the Antibacterial Drug Candidates

I. INTRODUCTION The resistance of pathogenic microorganisms to existing antibiotics is a major public health risk,1−4 and great efforts are currently being made to develop new therapeutic agents. (See, for instance, refs 5 and 6 and other works cited therein.) Recently,7−12 the title compound p-guanidinoethyl-phenol has been envisaged as a building brick for developing new antibacterial drugs. The efficiency of tetra-p-guanidinoethylcalix[4]arene (Cx1), which can be considered as a spatially organized tetramer of p-guanidinoethyl-phenol (mCx1) (Scheme 1), was found to be very promising against various Gram positive and Gram negative strains. Both Cx1 and mCx1 were studied with various membrane models, specifically phospholipid monolayers at the surface of water.13 The mCx1 monomer displayed a poor effect against neutral and negatively charged models, but the tetramer Cx1 imposed a strong perturbation of negatively charged membrane models, representative of bacterial membranes. PM-IRRAS investigations allowed us to propose a mechanism of action, which could © XXXX American Chemical Society

involve strong and synergistic interactions between the guanidinium groups and the membrane phosphate anions. Received: July 14, 2012 Revised: August 27, 2012

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dx.doi.org/10.1021/jp3069893 | J. Phys. Chem. A XXXX, XXX, XXX−XXX

The Journal of Physical Chemistry A

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The association of guanidine to calixarene macrocycles has not been deeply investigated, although some previous studies have reported very interesting contributions. Therefore, some works aimed at developing mimics of acetylcholine esterases,14,15 ligands for nucleotide recognition at the air− water interface,16 or cell transfection enhancers.17 Direct guanidination at the upper rim of calix[n]arenes, protected by ether functions at the lower rim, have allowed the design of DNA binders18 and, more recently, of artificial phosphodiesterases.19 para-[(Guanidino)methyl] calix[4]arenes integrating hydrophobic loops at the lower rim have been designed as protein oligomers stabilizers.20 Derivatives in which guanidine is attached to the calixarene core via a carbonyl function (with or without free −OH groups at the lower rim) have also been designed as reversible ligands of voltage-dependent potassium channels.21 Generally speaking, the use of guanidine organic derivatives as potential therapeutic agents is not new, and several compounds have deserved attention in the literature in relation to a variety of diseases.22 They can be considered as derivatives of the natural aminoacid arginine, whose side chain consists of a propyl chain attached to a guanidine group. Because the pKa of guanidinium is very high,23 guanidine is protonated in acidic, neutral, and even basic aqueous environments. The positive charge confers highly specific characteristics to guanidine compounds of medical interest, and careful analysis of the physicochemical properties of new drug candidates, such as the partition coefficient or the pKa values, is relevant to understand their activity and their mechanism of action. For instance, very recently, Salvio and coworkers19 have presented potentiometric evaluations of acidity constants of bis-1,2-, bis-1,3-, tris-, and tetra-para-guanidino calixarenes, protected at the lower rim by ether functions as well as their constitutive monomer, the N-(4methoxyphenyl) guanidine, in a DMSO/H2O 80:20 (v/v) medium and using Me4NOH as base. Their study has allowed the determination of the guanidinium pKa, which lies between 9.3 and 12.3, showing that a significant decrease in some acidities may be expected with respect to parent alkylguanidinium compounds. Predicting the pKa values of this type of systems is not trivial because large and opposite intramolecular and solvation effects are in competition. For instance, the different ionization states of mCx1 are represented in Scheme 2. The phenolate mCx1 structure 2 could be highly stabilized through intramolecular electrostatic interactions that are favored by (1) the negative charge delocalization on the phenol ring (Scheme 3) and (2) the flexibility of the ethylguanidinium group, which allows for the net charges getting close (see below). At this point, it may

Scheme 3. Mesomeric Forms of 2 Showing That Strong Stabilization of the Phenolate Charge by Electrostatic Interactions with the Guanidinium Group Is to Be Expected

be interesting to comment on the possible similarities of mCx1 with other systems. First, it is worth reminding that the pKa values of phenols are much lower than those of aliphatic alcohols due to the resonance stabilization of the negative charge, and for the same reason they are quite sensitive to substituent effects. For instance, substituents allowing to stabilize/delocalize further the negative phenolate charge will lower the OH group pKa; a prototypical example is 2aminophenol. In mCx1, the ethylguanidinium group does not allow for further delocalization, although eventually it can stabilize the negative charge through intramolecular Coulombic interactions. Second, the pKa of the phenol and guanidinium moieties in mCx1 might be compared with those of the equivalent groups in the aminoacids tyrosine and arginine, respectively. In the first case, however, the comparison is not as straightforward as in tyrosine; the final state is a dianion (phenolate + carboxylate), whereas in mCx1, it is a zwitterion (phenolate + guanidinium). In the case of arginine, the comparison makes sense because the pKa of the guanidinium group in the two systems will be potentially influenced by a negative charge (carboxylate and phenolate in arginine and mCx1, respectively). We emphasize that the third pKa of arginine is 12.48, therefore, lower than the pKa of alkylguanidiniums24 (13.3 for ethyl-guanidinium). Apart from the intramolecular effects, aqueous solvation effects have to be considered. In mCx1, they might possibly stabilize the acidic form 1 (total charge +1) or the totally deprotonated one 3 (total charge −1) to a larger extent than the monoprotonated forms 2 or 2′ (no net charge). Solvation does also control the equilibrium between these monoprotonated structures (zwitterionic and neutral, respectively). With the aim of better understanding the properties of antibacterial molecules based on mCx1 and of other compounds of pharmaceutical interest bearing a guanidine group, we report below a combined experimental and theoretical study of mCx1. Experimentally, the first pKa value of mCx1 has been determined in water medium by monitoring both pH and electric conductivity during a classical NaOH titration. The second one, corresponding, in principle, to the guanidine moiety, is expected to be ∼13, that is, is in the limit of classical titration methods in water. Both pKa values have then been calculated and analyzed with the help of quantum chemistry calculations using a discrete-continuum model of the solvent. The discussion focuses on the role of intramolecular and intermolecular interactions.

Scheme 2. Different Ionization States of p-Guanidinoethylphenol (mCx1)

II. EXPERIMENTAL AND COMPUTATIONAL DETAILS A. Chemicals. Volumetric solutions (HCl 0.1 M and NaOH 0.1 M) were purchased from Sigma-Aldrich (Sigma-Aldrich B



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(Vibrational frequencies in gas phase or solution have been used in each case.) These calculations have been done using the most stable structures found in our work, in parallel with many studies reported in the literature. Obviously, more elaborate free energy estimations would require a statistical treatment and a proper sampling of the conformational space. We have not envisaged such a treatment here that could be done using molecular dynamics simulations. Free energies in gas phase correspond to 1 atm and 298.15 K reference state, whereas free energies in solution correspond to 1 M at the same temperature. To calculate pK a values, one usually considers the equilibrium

Chemie, Steinheim, Germany). The mCx1 monomer was prepared as previously described.10 B. Apparatus. For pH determination, a pH-meter PHM210 (Radiometer Analytical SAS, Lyon, France) was used. The conductivity was evaluated using a conductivity-meter LF539 equipped with a cell Tetracon 96 from WTW (WTW, Weilheim, Germany). C. Theoretical Calculations. As a preliminary step, the potential energy surfaces for structures 1−3 in gas phase were explored at the B3LYP/6-31+G(d) level26 to locate the different energy minima. The use of diffuse functions was considered to be important here because they are essential for a realistic description of anions.27 The structures of phenol/ phenoxide, guanidine/guanidinium, and ethyl-guanidine/ethylguanidinium systems were also fully optimized at the same level. Calculations in aqueous solution were carried out for the same species using continuum and discrete-continuum models with the PCM approach.28−30 Simple continuum calculations are expected to provide qualitative trends for the investigated systems, and obtaining accurate calculations of solvation energies and pKa values requires, in general, the use of a combined discrete-continuum approach.31,32 In this case, the water molecules in the first solvation shell of the systems are explicitly included in the quantum chemical calculation. The number of water molecules in the first solvation shell cannot be unambigously determined and can change depending on protonation state. However, a total number of eight appeared to be appropriate in preliminary calculations. They correspond to the total number of solute−solvent hydrogen bonds that are likely to be formed: five for guanidinium or guanidine groups and three for the phenol group. This number was kept constant for all species (protonated or not) because error compensation (especially for entropic contributions) is optimized in this way.33 Likewise, the calculations for the phenol/phenoxide and guanidine/guanidinium calculations were done with three and five water molecules, respectively. Note that in discretecontinuum calculations there are often difficulties to select the most stable configuration of the complexes, and it is then compulsory to explore carefully the potential energy hypersurface. Full geometry optimization of the species in gas phase or solution at the B3LYP/6-31+G(d) level was followed by frequency calculations to confirm that the located structures correspond to energy minima. Only the most stable conformations found will be considered hereafter. Singlepoint energy calculations were performed on the most stable structures at the B3LYP/6-311+G(2df,2p) level in gas phase and solution. It is worth mentioning that a main limitation of DFT-based methods, in general, and of B3LYP, in particular, is the poor evaluation of dispersion energy, a limitation that can introduce significant errors in describing intramolecular interactions and hydrogen bonds with water molecules. To evaluate the error in the calculation of intramolecular interactions, some test calculations using the MP2/6-311+G(2df,2p) method were carried out. We assume that dispersion energy errors in hydrogen bonds roughly cancel out when total energy differences for different complexes are obtained. For comparison, MP2/6-311+G(2df,2p) energy calculations were also done to calculate the acidities of reference compounds (phenol, guanidine) in the gas phase. The calculations will be simply labeled B3LYP or MP2 below. Zero-point energy and thermal contributions to free energy were computed in gas phase and in solution at the B3LYP/6-31+G(d) level.

HA aq → A −aq + H+aq

(1)

although the following equations are sometimes used in the literature34,35 HA aq + OH−aq → H 2Oaq + A −aq

(2)

HA aq + H 2Oaq → H3O+aq + A −aq

(3)

When eq 1 is used, the pKa of HA is calculated from A− H+ HA ΔGaq = Gaq + Gaq − Gaq

pK a =

(4)

ΔGaq 2.303RT

(5)

where R is the gas constant, T is the temperature, and ΔGaq is the free energy of the ionization reaction. We have used expression 5 to calculate the absolute pKa of the reference compounds used in our work, that is, phenol and ethylguanidinium. We assume in that case the experimental Gibbs free energy of the solvated proton −270.28 kcal/mol (based on ideal gas approximation and solvation energy reported by Tissandier et al.;36 see the Supporting Information for details; an error of at least 2 kcal/mol is usually assumed37). We verified that eq 2 or 3 leads to similar results. The comparison with experimental data (see below) has allowed us to check the suitability of the methodology. The pKa values of mCx1 have been calculated using isodesmic reactions and experimental pKa data for the reference systems, in parallel with recent estimations of the gas-phase acidity of arginine38 HA aq + X −aq → A −aq + HX aq

pk a(HA) =

(6)

− 1 HX (GaqA − GaqAH − GaqX− + Gaq ) 2.303RT

+ pK a(HX)exp

(7)

where Gwaq is the free energy in aqueous solution for species W and HX is the reference species: phenol (for calculating the pKa of the −OH group in mCx1) or ethyl-guanidinium (for the calculation of the pKa of the guanidinium group). The experimental pKa values are 9.98 for phenol39,40 and 13.3 for ethyl-guanidinium.24 The interest for using expression 7 instead of a direct computation of absolute pKa (5) is two-fold: it avoids uncertainties related to the proton free energy and it leads to an excellent cancelation of computational errors ascribed to both theoretical level and solvation model, provided a convenient reference is chosen. A suitable choice needs to consider an acidic group that includes the main chemical characteristics of the target system in such a way that basis set C



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or correlation energy errors are comparable. (See our choice below.) All calculations were carried out with the program Gaussian 09.41 PCM calculations were done using the standard options of the program and included electrostatic and nonelectrostatic solvation energies. The optimized structures are provided in the Supporting Information.

although for pKa beyond 11 the glass electrode is no longer reliable due to alkaline error. During titration of a weak acid by the strong base NaOH, one observes two different zones, before and after the equivalence. Assuming a constant total volume (negligible NaOH volume added in front of the total one), the first zone corresponds to the neutralization of the weak acid AH, leading to the formation of the conjugated base A− and water. During titration, Na+ ions are accumulated in the solution, and a linear relationship between the solution total conductivity and the added NaOH volume is observed (Figure 1, linear regression line 1). When the point of equivalence is reached, the excess of OH− ions leads to a linear increase in the solution total conductivity (Figure 1, linear regression line 2) with a slope significantly different from line 1. The point of equivalence (Veq) is then determined at the intersections of the conductivity lines 1 and 2. To determine accurately the beginning of the weak acid titration (V0), a small amount of a strong acid (HCl 0.1 M) was added in the initial sample solution before titration. The pKa value was identified as the semiequivalent point of titration. According to this protocol, identification of the pKa of the phenolic group was achieved by simultaneously monitoring the pH and conductivity of a solution of 5.2 × 10−5 mol of mCx1 in 50.0 mL of water during its titration by 0.1 M NaOH (50 μL of HCl 0.1 M was added to the sample solution before titration). As shown in Figure 1, the determined pKa value is 9.2. Assuming that the pKa of the guanidine moiety is not accessible by classical titration in water (pKa > 13), this pKa has been attributed to the phenol part of mCx1 and has been confirmed (see hereafter) by theoretical calculations. B. Theoretical Results. Calculations on Reference Systems. The calculated gas-phase acidities of phenol and guanidinium are collected in Table 1; they show a very good agreement with available experimental data. When the MP2 method (which is much more computationally demanding) is used, the improvement of the results is not conclusive; therefore, in the following, only the DFT method will be employed as a good compromise between accuracy and calculation time. Free energies in solution and pKa of phenol and ethylguanidinium are shown in Table 2. (Ethyl-guanidine is used here instead of guanidine because its pKa will be used as a reference in eq 7 and it is structurally related to mCx1.) The pKa of ethyl-guanidinium is in perfect agreement with experiment, even when the simple continuum model is used. The pKa of phenol is a little underestimated in the discretecontinuum model, but the agreement can be considered as satisfactory because the difference is within the error assumed for the proton free energy of solvation (2 kcal/mol37 leading to 1.5 pKa units). The calculated pKa values could be improved by using parametric equations and data for different acidic functional groups equations, as proposed sometimes,42,43 but because we are mainly interested on relative acidities (see below), such a refinement was not considered necessary here. Note that the use of a simple continuum model is not appropriate in the case of phenol, which can be attributed to the importance of describing explicitly the first solvation shell in the case of anionic species (phenolate).32,44 In addition, radii used to construct the atomic spheres in the continuum model are difficult to define. (In general, solvent molecules are closer to the solute in the case of anions because of the strong electrostatic interactions, and this would suggest using smaller atomic spheres; however, anions are also characterized by a

III. RESULTS A. Experimental Determination. For weak acids with pKa values between 2 and 8, the direct observation of the titration

Figure 1. Experimental determination of the pKa of the phenolic part in mCx1.

Table 1. Calculated Gas-Phase Energies (kcal/mol) for Ionization of Phenol and Guanidinium at B3LYP and MP2 Levels at 298 K method

ΔEg

ΔHg

ΔGg

phenol

B3LYP MP2 exp

355.30 354.46

340.33 339.48 341.5446

guanidinium

B3LYP MP2 exp

245.04 246.06

347.83 347.00 351.445 347.547 347.9946 238.28 234.31 236.038 a

system

a

231.3 232.7

Using experimental data from Amekraz et al.48

Table 2. Ionization Free Energies (kcal/mol) and pKa of Phenol and Ethyl-Guanidinium in Aqueous Solutiona process phenol ethyl-guanidinium

continuum

discretecontinuum

exp

ΔG

pKa

ΔG

pKa

pKa

23.54 17.96

17.3 13.2

11.63 18.12

8.5 13.3

9.9839,40 13.324

a

Calculations were done using continuum and discrete-continuum models at the B3LYP level.

curve with a strong base like NaOH allows us to determine the point of equivalence with a good precision thanks to the presence of a very clear jump of pH. For very weak acids with pKa over 8, the jump of pH is poorly marked and becomes practically imperceptible for still higher pKa. In that case, a measure of the conductivity of the solution during the titration allows an accurate determination of the point of equivalence,25 D



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Figure 2. Optimized structures of mCx1 in the gas phase. Relative energies of conformers (ΔE = Eopen − Efold) and free energies (in parentheses) in kilocalories per mole. Calculations at the B3LYP level.

Table 4. Calculated and Experimental pKa values of mCx1a

Table 3. Energetics for Processes Connected to Gas-Phase Acidities of mCx1 at the B3LYP Level (kcal/mol) process

ΔE

ΔH

ΔG

1 → 2 + H+ 2 → 3 + H+ 1 → 2′ + H+ 2′ → 3 + H+

282.59 319.61 253.82 348.37

275.47 312.41 247.05 340.83

270.87 302.66 239.91 333.62

process

ref. acidity in calculations

1 → 2 + H+ 2 → 3 + H+

phenol/phenoxide guanidinium/guanidine

calculated

experimental

ΔpKa

pKa

pKa

−0.61 +0.92

9.3 14.2

9.2

a

Absolute pKa calculations were made using eq 7 and phenol and ethyl-guanidinium as reference compounds for the first and second pKa values, respectively. Calculations at the B3LYP level.

diffuse electron density that would be rather consistent with the use of larger atomic spheres.) Calculated mCx1 Acidities. The structures of mCx1 in its different protonation states have been investigated in gas phase. Each protonation state may display two conformations depending on the orientation of the guanidine/guanidinium group with respect to the phenyl ring, called folded and open hereafter. The optimized structures and relative energies in gas phase are presented in Figure 2. The results obtained show that in terms of potential energy, the folded structures are always more stable than the open ones. One might wonder whether dispersion contributions, which are poorly estimated by the B3LYP method, could further stabilize the folded structures.

Indeed, MP2 single-point energy computations on B3LYPoptimized geometries for the open and folded conformations of the neutral (nonzwitterionic) system PGH lead to a slightly higher preference for the folded structure. The open-folded energy difference increases from 0.22 to 1.37 kcal/mol. Hence, this energy increase (1.2 kcal/mol) can be considered as a rough estimation for the dispersion contribution. The analysis of other errors in B3LYP calculations would require a more elaborated study using highly correlated ab initio approaches, but this would be beyond the aim of this work. The energy difference is particularly large for the zwitterionic structure 2. This result is not surprising because in this case there are strong

Figure 3. Structures of mCx1 complexes with eight water molecules in solution and free energy differences (ΔG = Gopen − Gfold) in kilocalories per mole. Calculations at the B3LYP level. E



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Figure 4. Schematic representation of mCx1 and phenol acidities showing quite different values in gas phase but similar values in aqueous solution. (1 and phenol in gas phase are arbitrarily chosen as the free energy references for the comparison.)

electrostatic interactions between the phenolate and guanidinium moieties; in the folded structure, the guanidinium group is very close to the phenyl ring, one of the C(phenyl)···H(guanidinium) distances being as short as 2.2 Å. Thermal contributions reduce the energy differences for 1, 2, and 3 and reverse the open-folded stability in the case of 2′. Several gas-phase processes related to the acidities of mCx1 are presented in Table 3. (For simplicity, we report energies for only the most stable conformations.) Interesting comparisons can be made between these processes and the calculated gasphase acidities for phenol and guanidinium. For instance, the process 1→2+H+ requires a free energy of 271 kcal/mol, whereas the ionization process in phenol (Table 1) leads to 340 kcal/mol. The process 2→3+H+ requires roughly 303 kcal/mol, to be compared with 231 kcal/mol in the guanidinium ion (Table 1). From these results, one can estimate the influence of the intramolecular electrostatic interaction in the zwitterion 2 to ∼70 kcal/mol. Such a large energy effect is a direct consequence of the folding of the molecular structure, which becomes possible due the extra flexibility provided by the ethyl group. It may be also interesting to compare the −OH acidity in 2′ (process 2′→3+H+) with that of phenol: a free energy decrease of roughly 6 kcal/mol is predicted. Likewise, we can compare the −NH acidity in 1 (process 1→2′+H+) with respect to that of guanidinium: a free energy increase of ∼9 kcal/mol is predicted in this case. Finally, if we compare the processes 1→ 2+H+ (phenol ionization) and 1→2′+H+ (guanidinium deprotonation), then the second process is more favorable than the first one by as much as 31 kcal/mol, which corresponds to the energy difference between the tautomeric forms 2 and 2′, the latter being more stable in gas phase. The optimized structures (folded and open) for mCx1 solvated by eight water molecules in aqueous solution and the corresponding relative free energies are presented in Figure 3. Quite interestingly, the open structures are now systematically more stable than the folded ones. The most striking result concerns structure 2 because the open structure becomes more stable than the folded one despite its much lower stability in the gas phase (Figure 2). This stability inversion can be explained by the large dipole moment of the open structure of the

zwitterion 2 that leads to a greater interaction with the dielectric medium. The change from folded to open structures in aqueous solution is expected to have major consequences in terms of acidity because as noted above, the high gas-phase acidity is connected to the Coulombic intramolecular interactions in the folded geometries. Note that in all cases the open-folded energy difference is larger than the estimated error discussed above for the lack of dispersion energy (1.2 kcal/mol). Before discussing the acidities in aqueous solution, let us consider what is the most favorable structure of the monoprotonated species. In principle, one expects the zwitterionic structure 2 to be more favored in water, and this is indeed confirmed by the calculations. However, it is also important to estimate the energy difference to check whether 2′ plays a significant role on the chemical equilibria. Our calculations for the complexes with eight water molecules predict that the zwitterion 2 is more stable than the neutral structure 2′ by 6.6 kcal/mol in aqueous solution. This result, which contrasts with gas-phase data, suggests that 2′ can safely be neglected in the discussion of pKa values. Our calculations also show that using a discrete-continuum model is compulsory to describe correctly the relative stability of 2 and 2′. (If the simple continuum model is used, then the opposite relative stability is predicted.) Relative and absolute pKa values of mCx1 are calculated at the B3LYP level using eqs 6 and 7 and the most stable open structures in solution; the results are collected in Table 4. The first pKa in mCx1 (pKa1) corresponds to the −OH group, and the calculated value (9.3) is in excellent agreement with the experimentally measured value (9.2). It confirms that mCx1 (1) in aqueous solution exhibits only a slightly larger acidity than phenol (pKa = 9.98), despite its much higher gas-phase acidity (Table 3). This is of course a consequence of the relative solvation energies for the acid/conjugated base pairs 1/2 (q = +1, zwitterion, respectively) and phenol/phenoxide (q = 0, q = −1, respectively), as schematized in Figure 4, but behind this energy scheme, there is of course a main structural factor connected to the change from folded to open geometry in going from gas phase to water solution. F



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Finally, the second pKa (pKa2) of mCx1 corresponds to the deprotonation of the guanidinium group to yield the anion 3. This pKa is predicted to be ∼1 unit above that of ethylguanidinium (14.2 vs 13.3) and contrasts with the guanidinium pKa in arginine, which is significantly lower (12.48). Such a value corroborates the fact that it cannot be observed experimentally by usual titration techniques.

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IV. CONCLUSIONS The results presented above show that the first acidity of mCx1 corresponds to the phenolic group and is close to that of the unsubstituted phenol, although the pKa is a little smaller. The calculation details reveal that this happens despite huge differences existing between the gas-phase acidities of mCx1 with a protonated guanidine group (structure 1) and phenol. In fact, there is a large compensation between the intramolecular electrostatic interactions and the solvation effects on ionization energies, which separately amount to ∼70 kcal/mol. Overall, the predominant effect is the intramolecular electrostatic interaction in 2 between the phenolate moiety and the neighboring, positively charged, guanidinium group so that the zwitterionic monoprotonated intermediate is slightly stabilized with respect to either 1 or 3. A main characteristic of the investigated system is the possibility to change from the Coulombic-favored folded structures, more stable in gas phase, to open structures, which bear a very high-dipole moment and are more stable in aqueous solution. Considering the characteristic role of solvation energy in this system, one might wonder how the properties of mCx1 would evolve in transferring the molecule to nonaqueous media or to a water− hydrophobic interface, which is relevant to understand the antibacterial activity regarding the interaction with the bacterial wall. Work on this direction is under way.

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ASSOCIATED CONTENT

S Supporting Information *

Details of experimental free energy of the proton. Optimized geometry of all structures. Full ref 41. This material is available free of charge via the Internet at http://pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected], [email protected], [email protected]. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS We thank the French CINES for computational facilities (project lct2550). Financial support from the French ANR (CREAM project, ANR-09-BLAN-0180-01) and the Lorraine region (IMTS project) are gratefully acknowledged.

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REFERENCES

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Intramolecular Interactions versus Hydration Effects on p

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