α-amino acids like glycine (gly) (E Merck) and Dlalanine (ala) ,amino butyric acid (aba) and nor-valine (n-val) were used after drying as described earlier [15].

Ethylene glycol (LR, BDH) was purified by the usual method [16]. Ethylene glycol (LR, BDH) was refluxed with 2% - 3% NaOH (Merck) fpr 3 - 4 hours and then distilled; the distilled glycol was then dried over freshly baked anhy. Na₂SO₄ (Merck) for 4 - 5 days, then decanted off and fractionally distilled through a 2/3 m long vigreux column, rejecting the head and tail portions.

N,N-dimethyl formamide (DMF) (LR, BDH) was purified [16] first by distilling under reduced pressure in N₂ atmosphere and preserving the distillate over dry K₂CO₃ (Merck) for a week or so. The solvent was then decanted off and treated with pure P₂O₅ (Riedel) and finally distilled under reduced pressure.

The water content of the solvents were determined by Karl-Fisher titration and found to be less than 0.02-mol dm^–3 in each case.

Non-aqueous mixtures of co-solvent (ethylene glycol and N,N-dimethyl formamide) that have been used were 20, 40, 60, 80 and 100 wt% and were protected by storing in desiccators when not in use.

The solubility of these four amino acids were measured by the formol titrimetric method as described in our previous paper [12,17]. These measurements were taken at 15˚C, 20˚C, 25˚C, 30˚C and 35˚C temperatures. The low-cumhigh temperature thermostat used for all measurements was capable of registering temperatures having an accuracy of ±0.1˚C. Three sets of measurements were made for all the solutes by equilibrating the solutions from both above and below the required temperatures and at least two sets of measurements were made for all the solvents and the solubilities were found to agree to within ±1% to 1.5%.

The solvent parameters are listed in Table 1. The measured solubility (m) of the amino acids (on molal scale) is listed in Table 2.

As in the previous studies by Bates and coworkers on Tris [18] and by Kundu and coworkers [18,19] on nonelectrolyte like para-nitroaniline, benzoic acid and amino acids [15], glycine (G), diglycine (DG), and triglycine (TG), the Gibbs energies of solutions () of these amino acids on molal scale were calculated for each solvent using Eq.1.

where y is the molar activity coefficient of the solutes but taken tentatively to be unity in each solvent. True, since these amino acids are likely to be mostly in zwitterionic forms as in non-aqueous solvent mixtures [20, 21], the involved activity coefficient factor-RTlny in arising from interactions of dipolar solutes with large dipole moments may not be that small. But as there is neither the required experimental data nor any appropriate theoretical correlations for computing the same, these have been tacitly taken to be negligibly small, as is usually done for non-electrolytes [11]. This is because the effective contribution of activity coefficient factor –RTlny_s/y_R in the transfer free energetics = in particular which is our main concern likely to be hardly significant.

The free energies, at different temperatures are fitted by the method of least squares to an equation of the form;

where T is the temperature in Kelvin scale. The values of the coefficients a, b, c are presented in Table 3. These are found to reproduce the experimental data within ± 0.04 kJ·mol^–1. Transfer Gibbs energies, and entropies, of the amino acids from ethylene-glycol to N,N-dimethylformamide mixtures were calculated at 25˚C on mole fraction scale by using the following Eqs.3 & 4:

i.e.

and

here the subscript “s” and “R” refer to the co-solvent (ethylene glycol and N,N-dimethyl formamide) mixtures and reference solvent (ethylene glycol) respectively and M is the molar mass of the pure and mixed solvent. and values of amino acids thus obtained and presented in the Table 3. The estimated values shows an uncertainties in and are about ±0.05 kJ·mol^–1 and 2 kJ^–1·mol^–1, respectively.

Now (where X = G or S) may be ascribed as the sum of the following terms (assuming dipole induced dipole term to be negligibly small). i.e.

here, means for the transfer energy contribution of the cavity effect which is involved due to creation of cavities for the species in ethylene glycol and ethylene glycol and N,N-dimethyl formamide mixed solvent system and stands for the dipole-dipole interaction effect involving interaction between dipolar-zwitter-ionic amino acids and the solvent molecules, on the other hand, includes that for all other effects such as those arising from acid-base or shortrange dispersion interaction, solvophilic or solvophobic solvation and structural effects etc. Here values were computed by using Scaled particle theory (SPT) [17], assuming the solutes and solvent molecules as equivalent to hard-sphere models as dictated by their respective diameter (Vide Table 4).

and

were calculated by means of the Keesom-orientation expression [22] for in a solvent S, as given below

where

and

and that of as follows:

i.e., where N stands for Avogadro’s number, are the dipole moment of

solvents and amino acid molecules respectively (Table 4).

is the distance at which the attractive and repulsive interactions between the solvent and solute molecules are equal and is generally equal to where and are the hard sphere diameter of solvent and solute molecules respectively (Table 4) and α is the isothermal expansibility of the solvent and given by. Following Marcus [22] and Kim et al. [23] in order to get these term on mole fraction scale the quantity was again multiplied by the term.

which is the real mole fraction contribution due to dipole-dipole interaction [22]. Subtraction of and from the total we can get of amino acids. The values of, and are presented in Table 4.

The solubilities data reveals that the solubility of amino acids increases with the increase in temperature. Dl-α- amino butyric acid is somewhat more soluble in the ethylene glycol and N,N-dimethyl formamide solvent mixtures than Dl-alanine, which is contrary to the prediction based on the hydrophobic nature of these two compounds. This may be due to the relative law crystal lattice energy of Dl-α-amino butyric acid.

Figure 1 presents the variation of of amino acids with mole% N,N-dimethyl formamide in ethylene glycol and N,N-dimethyl formamide mixtures. Values of four amino acids (i.e. Glycine, DL-alanine, DL-α-amino butyric acid and DL-nor-valine) indicate their more or less gradual destabilizations with gradual increased concentration of N,N-dimethyl formamide in ethylene glycol and N,N-dimethyl formamide mixtures. As is composed of, and or others so their collective contribution to show such little complex nature of variation with mole% of N,N-dimethyl formamide.

The upward trends of profiles of all amino acids (Figure 2) indicate their relative destabilization with increased concentration of N,N-dimethyl formamide. The order of stability with respect to chemical contribution of solute-solvent interaction is Dl-val. > Dl-Aba. > Dl-Ala. > Gly.

The size of N,N-dimethyl formamide (0.498 Å) is greater than ethylene glycol (0.437 Å). Thus free energy change due to cavity formation is also more negative in N,N-dimethyl formamide relative to ethylene glycol. is more negative for the higher homologue among four amino acids having larger hard-sphere diameter (Table 4). On the other hand the dipole moment of N,N-dimethyl formamide (3.82D) is also greater than ethylene glycol (2.28D). Therefore, values are more negative in higher concentration of N,N-dimethyl formamide in this ethylene glycol and N,N-dimethyl formamide mixed solvent system. The order of is

Gly. > Dl-Ala. > Dl-Aba. > Dl-nor-val. As values are guided by dipole moment and hard-sphere diameter of both solutes (here amino acid) as well as solvent, the above order is well supported from Table 4.

values of the four amino acid represent the free energy change in the ethylene glycol and N,N-dimethyl formamide mixed solvent system due to different short range chemical interactions i.e. acid-base, dispersion, hard-soft, H-bonding, solvophilic/solvophobic interaction etc.

As the proportion of N,N-dimethyl formamide in the mixed solvent system will be gradually increased the solvent character may undergoes a gradual but material change in respect to the above types of chemical interactions.

While ethylene glycol is a good Bronsted acid, N,N-dimethyl formamide is not. Thereby anionic part (COO^–) of these four amino acids can be more solvated in ethylene glycol than N,N-dimethyl formamide due to acid-base interaction. Therefore with the increased concentration of N,N-dimethyl formamide values become more and more positive. On the other hand the cationic part () of the zwitterionic form of amino acids will be more solvated with the increased concentration of N,N-dimethyl formamide as it possess stronger lewis basicity and cationophilicity than ethylene glycol. Furthermore, in respect of H-bonding capacity ethylene glycol is more potential than N,N-dimethyl formamide. So amino acids will be less solvated with increased concentration of N,N-dimethyl formamide in ethylene glycol and N,N-dimethyl formamide mixtures.

It should be noted that N,N-dimethyl formamide (0.498 Å) is more polarisable than ethylene glycol (0.437 Å). Therefore N,N-dimethyl formamide, here undergo more stronger soft-soft and dispersion interactions with larger amino acids (i.e. α-aba & n-val).These phenomenon is well evidenced in the stability order i.e. (n-val.) > (α-aba.) > (ala.) > (gly.).

For larger amino acids like Dl-alanine, Dl-α-amino butyric acid and Dl-nor-valine along with soft-soft, dispersion and specific charge transfer interactions and another indirect and hence secondary interaction, namely solvophilic solvation (S_bS) is likely to be significant. Ethylene glycol, like H₂O, being extensively capable of intermolecular hydrogen bond formation, organizes a cage-like structure around organic moiety [here –CH₃ (Alanine), –CH₃–CH₃ (α-amino butyric acid), –CH₃– CH₃–CH₃ (non-valine)] by being induced by the latter, causing ‘solvophilic solvation’ which is similar to ‘hydrophobic hydration’ (H_bH) in aqueous solution.

Thus for larger amino acids although dispersion interaction is significant and tends to decrease values, the latter “solvophilic salvation” (S_bS) still decrease and tends to increases values with increase concentration of DMF as compared to those in pure ethylene glycol.

Thus the chemical contribution of transfer free energies, of these four homologous α-amino acids are guided by the composite effects of increased dispersion interaction, basicity and decreased acidity, hydrogen bonding effects and solvophobic solvation of ethylene glycol and N,N-dimethyl formamide mixtures as compared to that of reference solvent (ethylene glycol).

Figure 3 represents the change of these amino acids with ethylene glycol and N,N-dimethyl formamide mixtures. Here for all four amino acids rollercoaster type behavior are found. Like, can be taken to be composed of cavity, dipole-dipole and chemical interaction effects i.e.

where represents the difference of entropy change involved in creating appropriate cavities for accommodating the amino acids molecule in the reference solvent ethylene glycol and ethylene glycol and N,Ndimethyl formamide mixed co-solvent system in the present study.

stands for the dipole-dipole interactions originated due to dipolar amino acid molecule and dipolar mixed solvent system. term referred to as the chemical effect, stands for the structural interaction effect that appears due to the change of solvent structure induced by the amino acid molecules, if any, apart from that involved in the cavity effect. Now combined effect in values may represents such behavior as shown in Figure 3.

In Figure 4 Vs ethylene glycol and N,Ndimethyl formamide mixed solvent composition profiles is illustrated. For all amino acids regular increase of values with increase concentration of of N,Ndimethyl formadide is reflected. The value varies as Dl-n val. < Dl-α-aba. < Dl-ala. < gly. Now in order to understand the variation of for all four amino acids with increased DMF concentration in ethylene glycol and N,N-dimethyl formamide mixed solvent systems; one must note that solvation here occurs mainly through solvophobic solvation (S_bS), a phenomenon similar to hydrophobic hydration (H_bH) as in aqua-organic systems.

Solvation by this effect, solphobic solvation (S_bS) significantly decreases the entropy in ethylene glycol. Gradual desolvation with increased concentration of N,N-dimethyl formadide in ethylene glycol and N,Ndimethyl formamide mixtures for these amino acids is also guided by the increased hard-sphere diameter (Dl-n val. > Dl-α-aba. > Dl-ala. > gly.) of the amino acids.

Therefore as reflected in Figure 4, Dl-nor-valine will be more desolvated (i.e. lower) than other and glycine will be least desolvated (i.e. higher).

On the other hand observed monotonic increase of for these amino acids along with other interactions, is also guided by dispersion interaction. But solvophobic solvation being larger in the reference solvent, ethylene glycol than in the mixed solvents, it is quite likely that transfer of amino acids will disrupt the solvent structure, and hence make (amino acids) increasingly positive, as observed in Figure 4.