J. CHEM. SOC. FARADAY TRANS., 1990, 86(3), 501-505 50 1 Solution Studies of Haptens Containing Azobenzoate and Substituted Azobenzoate Anions in Water and in Methanol Angela F. Danil de Namor" and Rafic Traboulssi Department of Chemistry, University of Surrey, Guildford, Surrey GU2 5XH Mark Salomon US Army ET & DL (LABCOM), Power Source Division, Fort Monmouth, New Jersey 07703-5000, USA. The dissociation constants (PKd) of 0-, rn-and p-(4-hydroxyphenylazo)benzoic acid and chloro-substituted 4- hydroxyphenylazobenzoic acids in water at 298.15 K have been obtained by potentiometric titration. pK, values for azobenzoic acids are higher than those for unsubstituted benzoic acid, suggesting that the 4-hydroxyphenylazo group behaves as an electron-donating group.The dissociation Constants of chloro-substituted 4-hydroxylphenylazobenzoic acids are in excellent agreement with values calculated using the Hammett equation taking as reference PKd values for unsubstituted 4-hydroxyphenylazobenzoic acids and the substitution constants for chlorine substituents. Hammett substitution constants for 4-hydroxyphenylazo substit- uents are reported for the first time. Conductances, ion-pair formation constants of 0-, rn-and p-(Chydroxy- pheny1azo)benzoate and substituted azobenzoate salts in water and in methanol at 298.15 K are reported. Stokes radii for these anions in methanol and water are calculated. The data show that these anions are much more solvated in methanol than in water. Solubilities, solubility (ion activity) products and free energies of solution of a number of haptens in water and in methanol are reported.Free-energy data in water and in methanol are combined to calculate the transfer free energies of these electrolytes from water to methanol. A set of single-ion AG: values based on the Ph,AsPh,B convention for azobenzoate and substituted azobenzoate anions in the water-methanol solvent system is reported. Haptens are generally small molecules containing small acids, [SCl 2(pOHPhN,)BA] ; [6C1 2(pOHPhN,)BA], [4C1 groups known as antigenic determinants. These groups are 3(pOHPhN,)BA] and [2C1 4(pOHPhN,)BA] ; and their capable of combining specifically with antibodies. An under- sodium salts, [Cl(pOHPhN,)NaB]. standing of the interactions between an antibody with the It is well established that the hydroxyl group is among the hapten against which it is directed is of major importance in most common functional groups found in antibody molecules the field of biology, medicine and biochemistry.Relevant to and this group significantly contributes to hydrogen-bond this field of research is the study of interactions of haptens in formation in antibody-hapten complexation reactions. a reaction medium containing functional groups which may The main aim of this work is to determine transfer free be found in antibody molecules. An interesting group of energy, AG,", data for the haptens from water to methanol. haptens are those containing 0-,rn-and p-azobenzoate ions. These parameters are obtained from solution free energy, These haptens are known to interact with antibodies against AG:, data for the 1 : 1 electrolytes in water and in methanol.these ions. The degree of interaction is largely dependent on The calculation of AG: requires data for the ion-pair forma- the position of the carboxylate and azo group in the aromatic tion constants, K,, of these electrolytes in water and in meth- ring as well as on the presence of other substituents in the anol. This paper reports the following data. (a) The disso- 'p3molecule. ciation constants (pK,) of the acids obtained by poten- Therefore, solution thermodynamic data involving haptens tiometric titration. (b) Conductance data for their salts from in reaction media containing functional groups which are which K, values are derived.(c) Solution free energies of the known to be present in antibody molecules are useful param- haptens in water and in methanol as obtained from solubility eters in the interpretation of these complex systems. measurements of these haptens in these solvents. These data The haptens selected for this study are 0-,rn-and p-(4- are used for the calculation of the transfer free energies, AGP, hydroxypheny1azo)benzoic acid (p-OHPhN,) (fig. 1); their of the dissociated salts (M+ + X-) from water (H,O) to sodium salts, [(p-OHPhN,)NaB] ; the chloro-substituted methanol (MeOH). (d)A set of single-ion values for the trans- fer of anions from water to methanol based on the Ph,AsPh,B convention. Experimental 0-,rn-and p-(4-hydroxyphenylazo)benzoic acids and 2-chloro-4-, 4-chloro-3-, 6-chloro-2- and 5-chloro-2-(4-hydroxy- pheny1azo)benzoic acids were prepared from their respective aminobenzoic acids.The products were coupled with phenol. Finally, the highly coloured azobenzoic acids were precipi- tated at pH 2 and purified by recrystallisation from water and ethanol. The purity of the final products was checked by microanalysis carried out at the University of Surrey. The lc) melting points obtained are in good agreement with literature Fig. 1. Structures of (a)0-,(b)m-and (c) p(4hydroxyphenylazo)ben-data.3 Sodium salts were obtained by titrating the acid with zoic acids. NaHCO, until complete conversion had occurred. The solvent was evaporated and the salts were recrystallized from an aqueous solution of ethanol.The dissociation constants of the acids were determined by potentiometric titration with NaOH, using a TTT 80 titrator connected to a pHM82 standard pH meter. Continuous read- ings of pH and volumes added were recorded. The results of these measurements were analysed by using a MINIQUAD pr~grarn.~Conductance measurements and spectrophoto-metric titrations (UV)indicated that no additional species are formed in solution. It is only at very high pH that ionisation of the phenol group takes place. Conductance measurements were carried out at 25.00 & 0.01 "C in deionised water and in methanol (HPLC grade). The cell constant of the conductivity cell was deter- mined by the method described by Jones and Bradshaw.' The molar conductances of KCl were calculated using both Lind et aL6 and Fuoss-Hsia7 equations.The absolute ohm, the temperature scale and the molar mass of KC1 reported by Kay and coworkers' were used for these calculations. No sig-nificant differences were observed in the cell constant value derived from molar conductances of KCl obtained from the above equations. Thus, the cell constant value of 0.266 57 & 0.003 78 cm- ' (Lind et ~1.)~is in excellent agree- ment with that of 0.266 73 & 0.003 62 cm- ' (Fuo~s-Hsia).~ KCl (BDH, AnalaR) dried at 120 "C (concentration range 3 x 10-4-1.05 x mol dm-3) was used for these mea- surements. The stock solution of KCl was prepared from accurately weighed amounts of the purified salts dissolved in a known volume of deionised water.Conductance measurements for the haptens were deter- mined at various concentrations by successive additions of water or methanolic solutions of the salts. A stream of nitro-gen was passed through the solutions before measurements were carried out. For the solubility measurements, saturated solutions of the haptens were obtained by adding an excess of the solid to the solvent. The mixtures were left in a ther-mostat at 25.00 +_ 0.01 "C for several weeks. Samples were taken at the same temperature and analysed spectrophoto- metrically. The molar absorption coefficients of the anions in the two solvents (in units dm3 cm-' mol-') and the wavelengths of maximum absorption are as follows: o-(pOHPhN,)B-, 1.72 x lo4 at 348.1 nm (H,O) and 1.904 x lo4 at 354.4 nm (MeOH); rn-(pOHPhN,)B-, 1.712 x lo4 at 348.8 nm (H,O) and 2.997 x lo4 at 351.2 nm (MeOH); p-(pOHPhN,)B-, 2.16 x lo4 at 355 nm (H,O); 3.02 x lo4 at 359.8 nm (MeOH); 5C1 2-(pOHPhN,)B-, 1.554 x lo4 at 354.4 nm (H,O) and 2.208 x lo4 at 358.2 nm (MeOH); 6C1 2-(pOHPhN,)B-, 2.17 x lo4 at 353.5 nm (H,O) and 2.556 x lo4 at 358.9 nm (MeOH); 2C1 4-(pOHPhN,)B-; 2.52 x lo4 at 355.2 nm (H,O); 1.198 x lo4 at 359.5 nm (MeOH); 4C1 3-(pOHPhN,)B-, 3.760 x lo4 dm3 at 354.4 nm (H,O) and 2.354 x lo4 at 360 nm (MeOH).For solubility measurements solvate formation was tested by placing small amounts of the acids and salts used in closed vessels over the appropriate solvent at 298.15 K.The acids or salts were weighed from time to time in order to detect any uptake of the solvent. Solvate formation was not detected. All results are expressed in terms of the defined calorie? for the isothermal process at 298.15 K. Results and Discussion Dissociation Constants of the Acids The extent to which acids (HA) are dissociated is currently expressed in terms of the dissociation constant of the acid, Kd t 1 cal = 4.184 J. J. CHEM. SOC. FARADAY TRANS., 1990, VOL. 86 which refers to the process as described by: AH + H,OeA-+ H30+. (1) In table 1, values for the dissociation constants (expressed as pKd = -log Kd) for the azobenzoic and substituted azoben- zoic acids in water at 298.15 K as obtained from poten- tiometric titrations are given.The standard deviations of the data are also included in table 1. Attempts were made to obtain pKd values for these acids from conductance measurements. The results although not accurate enough to be reported (mainly due to their relatively low solubility in water) were analysed by the method described by Salomon' elsewhere and were found to differ by not more than 0.5 log units from the data given in table 1. In order to evaluate the effect of the pOHPhN, substituent group, the pKd value for benzoic is included in table 1. The ionisation of substituted benzoic acids in water was the model used by Hammett'4-'6 and by Burckhard and to establish the electron donating or electron withdrawing properties of substituent groups.'' The introduction of a substituent on the phenyl group will result (depending on the nature of the substituent) in an increase or decrease of the ionisation of the acid which will be reflected in the pKd values. Thus an increase in acidity (lower pKd values) is expected to be observed when the substituent group is electron withdrawing and a decrease (higher pKd values) for an electron donating substituent. It is clear from the results shown in table 1 that the introduction of the 4-hydroxyphenylazo (pOHPhN,) group in the structure of benzoic acid has an electron-donating mesomeric effect and therefore, azobenzoic acids are weaker (higher pKd values) than benzoic acid in water. This effect is greater for o-(pOHPhN,)BA and lower for p-(pOHPhN,)BA.The Hammett substituent constants (a*)defined" as: a* = pKd(BA) -pKd[(pOHPhN,)BA] calculated from data given in table 1 are -1.1 1, -0.88 and -0.36 for 0-,rn-and p-(pOHPhN,)BA, respectively. One of the most interesting features of these results emerges when these constants (a*)are compared with those reported in the literature for azophenyl substituents, (PhN,)" (a* = +0.29 and 0: = 0.32) since an opposite effect (electron withdrawing) is observed. The pKd values observed for the chlorosubstituted azoben- zoic acids are in excellent agreement with the calculated values obtained from a rearrangement of eqn (2), taking as reference the appropriate unsubstituted 4-hydroxyphenylazo benzoic acid and the corresponding G* values given in the literature for chlorine substituents (a; = 0.37, a: = 0.24 and a,*z 1.28).'9*20 Thus pKd[(pOHPhN,)BA] -o*(chlorine) = pKd[Cl(pOHPhN,)].(3) Using eqn (2) a* values for the chloro substituted 4-hydroxyphenylazobenzoic acids were calculated. The results are as follows: 5C1 2-(pOHPhN,)BA, -0.73; 6C1 2-(pOHPhN,)BA, 0.27; 2C1 4-(pOHPhN,)BA, 0.62 and 4C1 3-(pOHPhN,)BA, -0.43. Ion Association Constants of Hapten Salts in Water and in Methanol at 298.15 K Data for molar limiting conductances, A" and the association constants for the hapten salts in water and in methanol at 298.15 K are listed in table 2. Also included in this table are the number of conductance measurements carried out and the range of concentrations used for these measurements.Conductance data at the various concentrations were J. CHEM. SOC. FARADAY TRANS., 1990, VOL. 86 Table 1. pK, values of azobenzoic, substituted azobenzoic and K, values for the hapten salts in water and in methanol are related acids in water at 298.15 K relatively small and therefore, these salts are considered to be highly dissociated in water and in methanol. Using literature acid PK," PK,' data for the ionic molar conductances (2:) for sodium in o-(pOHPhN,)BA 5.31 f0.10 water22 and in methanol,23 corresponding data for the m+O HPhN ,)B A 5.08 k0.08 anions (A?) in these solvents are calculated. Details are given p-(pOHPhN,)BA 4.55 f0.08 in table 3. 5C12-(pOHPhN ,)BA 4.93 f 0.07 4.94 These data are used to calculate Stokes law radii (rJ.For6C12-(pOHPhN ,)BA 3.93 f0.06 4.03 2C14-(pOHPhN ,)BA 3.58 f0.04 3.27 these purposes, the following expression is used: 4C13-(pOHPhN ,)BA 4.63 f0.08 4.84 rs = Fz/6nqN2,BA 4.20' Values for the viscosities (q)of water and methanol are those This work. 'Calculated values, see text. 'Ref. (lOH13). given in the literature.2' Stokes radii for these anions in water and in methanol at 298.15 K are also included in table analysed by the Fuoss-Hsia equation using the expansion 3. With a few exceptions [rn-(pOHPhN,)B- and perhaps 4C1 given by Fernandez Prini2' This equation was solved by a 3-(pOHPhN2)B-], the Stokes radii are close to the values for pit mapping method' in which the square of the standard the benzoate anion (2.85 A) and for the o-chlorobenzoate error, s is minimised; s is defined by: (3.05 A) and rn-chlorobenzoate (2.97 A) anions.24 Comparison between the r, values in water and in methanol shows that (4) the rs values for these anions in methanol are almost twice For three degrees of freedom, the standard error of estimate the values observed in water, which suggests that these for the molar conductance is simply anions may be much better solvated in methanol than in water.If this is the case, these anions are to be characterised (5) by negative single-ion transfer free energies values from water Although it is shown by the uAvalues that the present theo- to methanol. These data are obtained from solubility mea- retical treatment does not fit the results so precisely as surements of these electrolytes in water and in methanol as expected one has to consider (a)the complexity of the system detailed below. under study and (b)in the context of this work, K, values are Solubilities, Free Energies of Solution in Water and in ancillary data required for the calculation of the free energy of solution AG,".We have shown in a previous publicationz5 Methanol.Derived Free Energies of Transfer that variations in K, do not lead to significant changes in the Solubility data for fifteen haptens in water at 298.15 K are AG," values. Therefore, we will argue that the K, values reported in table 4. Since the solution free energies, AG,", to reported in table 2 are good enough for the purpose of this be calculated are the values corresponding to the fully disso- work.ciated acid (H+ + X-) or salt (M' + X-), corrections for ion Table 2. Limiting conductivities, A', in association constants, K, ,of hapten salts in water and in methanol at 298.1 5 K no. of concentration haptens measurements range/mol dm - Ao/cm2Q-' mol-' Ka CA water o-(pOHPhN ,)NaB 17 5 x 10-4-3.1 x 85.21 9.49 0.36 m-(pOHPhN ,)NaB 14 1.8 x 10-5-3.0 x 72.65 1.79 x 10, 0.57 p-(pOHPhN ,)NaB 11 6.0 x 1OP4-9.8 x lop4 86.16 54.99 0.33 5C12(pOHPhN2)NaB 11 1.3 x 1OP4-5.0 x 81.45 8.28 2.25 6C12( pOH PhN JNaB 2C14(pOHPhN JNaB 9 12 2.0 x 10-4-5.0 2.3 x 10-'-2.7 x x 81.05 87.67 10.67 95.80 0.94 1.25 4C13(pOHPhN2)NaB 15 0.6 x 1OP4-9.6 x lop4 methanol 77.96 3.46 0.53 o-@OHPhN ,)NaB 15 (0.71-12.56) x 77.36 1.82 0.28 m-(pOHPhN ,)NaB p-(pOHPhN ,)NaB 17 10 (0.54-7.56) x (1.58-14.1) x lo-" 81.10 71.55 0 0.47 0.54 0.45 5C12(pOHPhN2)NaB 15 (1.21-13.17) x 78.69 11.16 0.15 6C12(pOHPhN ,)NaB 13 (2.49-13.54) x 75.23 0 0.15 2ClqpOHPhN ,)NaB 15 (1.27-14.22) x 65.91 0 0.19 4C13(pOHPhN2)NaB 14 (1.31-14.93) x lop4 73.88 7.68 0.26 Table 3.Ionic limiting conductivities and Stokes radii for azobenzoate and substituted azobenzoate anions in water and in methanol at 298.15 K anion Io"/cm2Q-' mol-' (H,O) r$A" ' Ilo'/cm2 Q-' mol-' (MeOH) r$A" ' o-(pOHPhN,)B -35.11 2.62 30.36 4.96 rn-(pOH PhN ,)B -22.55 4.09 34.10 4.42 p-(pOHPhN ,)B -36.06 2.56 24.55 6.14 5C12(pOHPhN ,)B -31.35 2.94 3 1.69 4.75 6C12(pOHPhN2)B-30.97 2.98 28.23 5.34 2C14(pOHPhN2)B-37.57 2.45 18.91 7.97 4C13(pOHPhN ,)B -27.86 3.31 26.88 5.61 Calculated from Ao values given in table 2 and A:,+ in water given in ref.(22). 'See text. Calculated from Ao values given in table 2 and I&+ in methanol given in ref. (23). J. CHEM. SOC. FARADAY TRANS., 1990, VOL. 86 Table 4. Solubilities and free energies of solution (molar scale) of 1 : 1 haptens in water at 298.15 K haptens o-(pOH PhN JBA m-(pOHPhN,)BA P-(pOHPhN,)BA 5C12(pOHPhN ,)B A 6C12(pOHPhN2)BA 2C14(pOHPhN2)BA 3Cl(pOHPhN,)BA o-(pOHPhN ,)NaB m-(pOHPhN,)NaB p+OH PhN ,)NaB 5C12(p0HPhN2)NaB 6C12(pOHPhN2)NaB 2C14(pOHPhN2)NaB 4C13(pOHPhN2)NaB solubility/mol dmP3 Ka PK," AG,"/cal mol -' 4.67 x 10-~ 2.04 x 105" 8.69 11 847 1.41 x 10-4 1.20 x 105" 9.04 12 326 4.16 x 10-5 3.55 x 104" 9.28 12 660 1.39 x 10-4 8.51 x lo4" 8.91 12 159 1.01 x 10-3 8.51 x 103 7.08 9 652 2.40 x 10-3 3.80 x 1030 6.35 8 658 8.33 x 10-5 4.27 x 104" 8.94 12 193 1.07 x lo-' 9.45' 2.47 3 370 6.06 x lo-, 1.72 x lo-' 1.8 x 55.P 3.62 3.98 4 939 5 430 2.16 x lo-, 8.P 3.55 4 843 3.04 x lo-' 1 1.Ob 1.87 2 251 1.24 x lo-' 96.p 4.3 1 5 880 2.37 x lo-, 3.5' 3.42 4 665 " From pK, values for the acids given in table 1.Values given in table 2 rounded off to the next highest or lowest figure. Table 5. Solubilities and free energies of solution (molar scale) of 1 : 1 haptens in methanol at 298.15 K solubility/ haptens mol dmP3 K" pK," AG,"/cal mol-' o-(pOHPhN,)NaB 5.29 x lop2 2.0 3.19 4352 m+OHPhN,)NaB 5.98 x lo-, 0 3.10 4320 p-(pOHPhN,)NaB 4.02 x lo-, 0.5 3.36 4582 5C12(pOHPhN2)NaB 1.33 x lo-' 11.0 4.16 5672 6C12(pOHPhN2)NaB 7.29 x lo-, 0 2.98 4058 2C14(pOHPhN2)NaB 2.098 x lo-' 0 2.33 3172 4C13(pOHPhN2)NaB 2.17 x lo-' 7.7 3.81 5196 ~~ ~ " Values given in table 2 rounded off to the next highest or lowest figure.Table 6. Free energies for the transfer of 1 : 1 haptens and for the anions (Ph4AsPh4B convention) from water to methanol at 298.15 K in kcal mol -AG,O'(M+ + X-) AG; '(X-) haptens (H,O -+ MeOH) (H,O -+ MeOH) o-(pOH PhN ,)NaB 0.98 -1.07 rn-(pOH PhN ')NaB -0.62 -2.67 p-(pOHPhN ,)NaB -0.85 -2.90 5C12(pOH PhN JNaB 0.83 -1.22 6C12(pOHPhN2)NaB 1.51 -0.54 2C14(pOHPhN2)NaB -2.71 -4.76 4C13(pOHPhN2)NaB 0.53 -1.53 "From AG," in water and in methanol given in tables 4 and 5.'By subtracting AG;(Na+)(H,O -+ MeOH) = 2.05 kcal mol-' [ref. (26)] from AG;(M+ + X-)(H,O --t MeOH) values given in this table. Table 7. Transfer free energies of the dissociated azobenzoic and substituted azobenzoic acids from water to methanol at 298.15 K in kcal mol- ' AGF[H+ + X-1" AG,"= [H+ + X-]' acid (H,O + MeOH) (MeOH) o-(pOHPhN,)BA 1.42 13.27 m-(pOHPhN,)BA -0.18 12.15 p(pOHPhN,)BA -0.4 1 12.25 5C12-(pOHPhN2)BA 1.27 13.43 6C12-(pOHPhN JBA 1.95 11.60 2C14-(pOHPhN2)BA 2.27 10.93 4C13-(pOHPhN ,)BA 0.97 13.16 "From single-ion values for the anions given in table 6 and AG,O[H+](H,O +MeOH) = 2.49 kcal mol-' based on the Ph4AsPh4B convention given in ref.(28). 'From AG,"[H++ X-](H,O + MeOH) (column 2) and AG,"[H+ + X-](H,O) given in table 4. association are made in order to calculate the ionic concen- trations, ci . For this purpose, K, values for the azobenzoic and substituted azobenzoic acids were derived from pK, values given in table 1. For the salts, K, values in water are those given in table 2. The solubility (ion activity) product, K,",is calculated by: K,"= C~Y: (7) where yrt is the mean molar ionic activity coefficient. With the lack of a better approximation, the extended Debye- Hiickel equation was used to calculate the yk values.For the ion size parameter, a', values between 5.0 and 6.5 A were used for the calculation of y* . It must be emphasized that y* values are not affected to any significant extent by ion size25.2 6 parameter (ao)changes. The solubility (ion activity) product, reported as pK," and the standard Gibbs free energy of solution, AG," for the dissociated acids (H+ + X-) and salts (M' + X-)are given in table 4. Solubility data for the hapten salts in methanol are listed in table 5. Also given in this table are the pK," and AG," values for these salts in meth- anol at 298.15 K. K, values used for these calculations are those from table 2. In order to eliminate inequalities attrib- uted to the contribution of the crystal lattice energy, transfer free energies of the dissociated electrolytes from water to methanol are reported in table 6.These data refer to the process described by : M+(H20)+ X-(H20) -+ M+(MeOH)+ X-(MeOH) (8) which involves the transfer among these two solvents in their pure state. The Ph,AsPh,B convention27 is used for the cal- culations of the single-ion AG," of a number of anions from water to methanol. The single-ion AG," value for the sodium cation used for these calculations is that given in the liter- ature.26 Large variations are observed among the AG," values for the different anions. In all cases, AG," values are negative indi- cating that these anions are better solvated in methanol than in water. These data are now compared with corresponding single-ion AG," values from water to methanol based on the Ph,AsPh,B convention reported in the literature2* of related anions such as: benzoate (B-), AG," = 1.77 kcal mol-'; p-nitrobenzoate (pN02B-), AG: = 1.07 kcal mol-' and p-methylbenzoate (pMeB-), AG," = 0.96 kcal mol-', in order to see the effect of the substituent group on the AG," values.The results are in line with the observation that an increase in the size of the anion will lead to more negative AG," values, an effect which was attributed to the non-electrostatic contribu- tion to the overall process of solvation.28 However, we think J. CHEM. SOC. FARADAY TRANS., 1990, VOL. 86 505 that it is rather dangerous to give an interpretation of the process on the basis of free energy data without considering the contributions made by the transfer enthalpy, AH," and entropy, AS: to the AG," values.Heats of solutions of these electrolytes in water and in methanol are being carried It is of interest to emphasize that except for 9 10 11 12 M. Salomon, J. Solution Chem., 1986, 15, 237, see also G. Beach, Fortran IV in Chemistry (John Wiley, London, 1975). F. C. Brockman and M. Kilpatrick, J. Am. Chem. SOC.,1934,56, 1483. B. Saxton and H. F. Meier, J. Am. Chem. SOC., 1934,5fi, 1918. F. C. Dippy, F. R. Williams and R. H. Lewis, J. Chem. Soc., 1935,343. rn-(pOHPhN,)B -a reasonable correlation is found between the single-ion AG: values (table 6) and the differences between the values for the Stokes radii in methanol and water (table 3).Combination of AG," values for these anions with AG,"[H+](H,O + MeOH) based on the Ph,AsPh,B conven-tion, yields the transfer Gibbs free energies of the dissociated 13 14 15 16 17 18 F. C. Dippy and F. R. Williams, J. Am. Chem. SOC.,1934, 1888. L. P. Hammett, Physical Organic Chemistry (McGraw Hill, New York, 2nd edn, 1970). L. P. Hammett, J. Chem. Educ., 1966,43,464. L. P. Hammett, Chem. Rev., 1935,17,225. G. N. Burckhardt, Nature (London), 1935,136,684. G. N. Burckhardt, W. G. K. Ford and E. Singleton, J. Chem. azobenzoic and substituted azobenzoic acids from water to Educ., 1936, 17. methanol (table 7).Since AG,"(H+ + B-) values in water are known (table 4), corresponding AG," values in methanol are calculated and the data are also listed in table 7.19 20 M. S. Isaacs, Physical Organic Chemistry (Longman Group UK Ltd, 1987). D. Perin, in pKa Prediction for Organic Acids and Bases (Chapman and Hall, London, 1981). R. T. gratefully acknowledges the scholarship given by The 21 R. Fernandez Prini, Physical Chemistry of Organic Solvent Systems, ed. A. K. Covington and T. Dickinson (Plenum Press, Hariri Foundation. London, 1973). (Butterworths, 2nd edn, 1973). References 23 R. L. Kay, C. Zawoyski and D. F. Evans, J. Phys. Chem., 1965, 69,4208. 1 L. Pauling and D. Pressman, J. Am. Chem. SOC.,1945,67, 1003. 2 D. Pressman, A. B. Pardee and L. Pauling, J. Am. Chem. SOC., 1945,67, 1602. 3 D. Pressman, M. Siege1 and L. A. R. Hall, J. Am. Chem. Soc., 1945,76,6336. 4 A. Sabatini, A. Vacca and P. Gans, Talanta, 1974,21, 53. 5 G. Jones and B. C. Bradshaw, J. Am. Chem. Soc., 1933,551780. 6 J. E. Lind, J. J. Zwolenik and R. M. Fuoss, J. Chem. SOC., 1959, 81, 1557. 7 R. M. Fuoss and K. L. Hsia, Proc. Natl. Acad. Sci., 1967, 57, 24 25 26 27 28 29 G. Milazzo, Electrochemistry (Elvesier, Amsterdam, 1963). A. F. Danil de Namor, T. Hill and E. E. Sigstad, J. Chem. SOC., Faraday Trans. I, 1983,79,2713. A. F. Danil de Namor, E. Contreras and E. E. Sigstad, J. Chem. Soc., Faraday Trans. I, 1983,79, 1001. B. G. Cox, G. R. Hedwig, A. J. Parker and D. W. Watts, Aust. J. Chem., 1974,27,477. M. H. Abraham, T. Hill, H. C. Ling, R. A. Schulz and R. A. C. Watt, J. Chem. SOC.,Faraday Trans. 1, 1984,80,489. A. F. Danil de Namor and R. Traboulssi, work in progress. 1550. 8 Y. C. Wu, W. F. Koch, W. J. Hamer and R. L. Kay, J. Solution Chem., 1987,16,985. Paper 9/02806D; Received 3rd July, 1989