Hydrophobic ion pairing

This section describes the nucleophilic reactions—acyl transfer reactions mostly—promoted by micelles and polysoaps. The nucleophiles are imidazoles, oxyanions and thiols, the same catalytic groups found ubiquitously in the enzyme active site. These nucleophiles are remarkably activated in the anionic form in the presence of cationic micelles and cationic polysoaps. These results are explained by the concept of the hydrophobic ion pair (Kunitake et al.,... 

In previous sections, numerous examples of anion activation by cationic micelles and polysoaps were presented. The extent of rate augmentation— 102—lO -fold—cannot be rationalized in terms of concentration effects alone. We believe that these observations are explained most reasonably by the concept of the hydrophobic ion pair (Kunitake et al., 1976a). According to this concept, anionic reagents are activated probably due to desolvation when they form ion pairs with an ammonium moiety in a hydrophobic microenvironment. The activation of anionic species in the cationic micellar phase... 

The ease of formation of hydrophobic ion pairs, and hence the rate acceleration, will be determined by the hydrophobic and electrostatic interactions between the anionic and cationic species. Lapinte and Viout (1974) found that the nucleophilic order OH- > CN > C6H50- in water was completely reversed in CTAB micelles hydrophobic phenoxide ion is activated better by the micelle. The micellar binding of phenols and phenoxides was determined by Bunton and Sepulveda (1979). Similarly, hydrophobic hydroxamates are activated much better than their hydrophilic counterparts. In the same vein, the extent of activation correlates approximately with the hydrophobic nature of aqueous aggregates as estimated by Amax of methyl orange (Table 7) and of picrate ion (Bougoin et al., 1975 Shinkai et al., 1978f Table 5). 

The formation of hydrophobic ion pairs may be envisaged as in Fig. 6. This may be considered as a microscopic counterpart of phase transfer catalysis. 

Fig. 6 Formation of a hydrophobic ion pair a comparison with phase transfer catalysis...

As an extension of the research on hydrophobic ion pairs, ion pairs such as tetraethylammonium hydroxamate [63] have been prepared and their nucleophilic reactivity estimated in organic solvents (Shinkai and Kunitake, 1976d Shinkai et al., 1979a). The ion pair showed very high nucleophilicity toward... 

As mentioned repeatedly, a variety of anionic reagents are highly activated in the hydrophobic microenvironment of cationic micelles and polysoaps. The range of anionic reagents studied in the past includes imidazole, hydroxide, thiolates, oximates, hydroxamates, carboxylates and carbanions. Polyanionic coenzymes are similarly activated. These results can be interpreted in a unified way by the concept of hydrophobic ion pairs, and the major source of activation seems to be concentration and desolvation of the anionic reagent in the... 

Meyer, J.D., J.E. Matsnnra, J.A. Rnth, E. Shelter, S.T. Patel, J. Bansch, E. McGonigle, and M.C. Manning, Selective precipitation of interleukin-4 using hydrophobic ion pairing a method for improved analysis of proteins formulated with large excesses of human serum albumin. Pharm Res, 1994. 11(10) 1492-5. 

Because of their ionic nature, most synthetic colors require ion-pair reagents. When a hydrophobic ion of opposite charge (counter ion) is added, hydrophobic ion pairs are formed, and hence the retention of the ionized substance on a nonpolar reverse phase is enhanced to achieve higher selectivity and peak symmetry (135,168,188). 

If polar sequences or short phosphopeptides fail to bind to the stationary phase upon loading, the acetonitrile content of the eluent could be reduced to 3%. A further decrease is not possible for standard Ci8-columns, as the stationary phase could collapse reducing the loading capacity and the reproducibility. Instead, TFA could be replaced by the more hydrophobic ion-pair reagent HFBA (heptafluorobutyric acid), or a stationary phase with a special coating compatible with pure aqueous eluents could be used (e.g., Aqua-column from Phenomenex). 

Thomlinson [78] was the first chromatographer to point out that the classical electrostatic ion-pair concept did not hold for IPRs that were usually bulky hydrophobic ions he also emphasized that in the interfacial region between the mobile and the stationary phases, the dielectric constant of the medium is far lower than that of the aqueous phase. Chaotropes that break the water structure around them and lipophilic ions that produce cages around their alkyl chains, thereby disturbing the ordinary water structnre, are both amenable to hydrophobic ion-pairing since they are both scarcely hydrated. The practical proof of such ion-pairing mode can be found in References 80 and 81 many examples of such pairing modes are reported in the literature [79-86],... 

It was also more interesting to observe that the valne of hydrophobic ion-pairing equilibrium constant runs counter to predictions of a purely electrostatic approach since it increases with increasing size of the pairing ion [107,109,112] for both organic and... 

The chromatographic estimate of ion-pairing equilibrium constant via IPC [87,114] will be thoroughly detailed in Chapter 3 (Section 3.1.2) and will confirm that separative techniques are particularly valuable for ascertaining the nature of hydrophobic ion-pairing. 

Section 2.5.3 in Chapter 2 expounded upon the hydrophobic ion-pair concept The peculiarities of this association mode, not even likely in the Bjerrum s model, were elucidated. Electrostatic attraction is only part of the story and solvophobic interactions are crucial to rationalize experimental evidence that often runs counter to the pristine electrostatic description of the process. 

Interestingly, the estimates of pairing constants obtained by the fitting of retention equations to experimental data increase with increasing analyte chain length (lipophilicity) [60] thereby supporting the hydrophobic ion-pairing concept. 

The model was recently tested to determine whether it was able to model analyte retention in the presence of novel and unusual IPRs (see Chapter 7) such as chaotro-pic salts and ionic liquids. Chaotropes that break the water structure around them and lipophilic ions (classical IPRs and also ionic liquids) that produce cages around their alkyl chains, thereby disturbing the ordinary water structure, are both inclined to hydrophobic ion-pairing since both are scarcely hydrated. This explains the success of the theory, that is predictive in its own right, when neoteric IPRs are used [64]. Recently a stoichiometric model (vide supra) was put forward to describe retention of analytes in the presence of chaotropic IPRs in eluents [18] but its description of the system is not adequate [64]. 

While a hydrophobic ion-pair is retained on hydrophobic stationary phases better than an ionized analyte, the retention of the duplex on normal phases is easily predicted to be lower than that of the ionized analyte because polar interactions are reduced. Actually the trend of k versus IPR concentration under normal phase IPC is the opposite of reversed phase IPC [34]. An aminopropyl, a cyanoethyl, and a silica stationary phase were compared for the analysis of alcohol denaturants. The cyanoethyl phase was selected and anionic IPRs were used to reduce retention of cationic analyte, suppressing their interactions with negatively charged silanols... 

Even if anionic chaotropes are the most popular neoteric IPRs, polarizable cations such as sulfonium and phosphonium reagents showed single selectivity toward polarizable anions their behavior was rationalized on the basis of their chaotropicity. Probe anion retention generally increases in the order of tributylsulfonium < tetrabutylammonium < tetrabutylphosphonium. Interestingly, retention was found to be influenced by the kosmotropic/chaotropic character of both the IPR and the probe anion [93] and this confirms the peculiarities of hydrophobic ion-pairing. Quaternary phosphonium salts provided increased selectivity compared to ammonium in the IPC of heavy metal complexes of unithiol [112]. 

The use of chaotropic counteranions for a chromatographic separation is beneficial as a method development strategy. These modiflers may replace the need for changing column type and/or addition of hydrophobic ion-pairing reagents for the more challenging separations. Further studies are needed to fully elucidate the detailed mechanism of chaotropic mobile-phase additives. 

Figure 7.23 Hydrophobic ion pairing of cytochrome c (Cc) with fluorinated surfactants KDP or Krytox (A) Dark aqueous solution of the haem protein, Cc. (B) Krytox dissolved in PFMC. (C) A biphasic system is initially observed with Cc in the aqueous (top) phase. (D) On stirring, Cc is extracted into the lower fluorous phase as it forms ion pairs with Krytox molecules. (E) If Krytox alcohol (no acidic group) is used, ion pairing is not possible and Cc stays in the aqueous phase. Note Cc and Krytox molecules are not drawn to scale. HIP complexes with only one Cc molecule surrounded by Krytox molecules are shown for clarity. [Reprinted with permission from Angew. Chem. Int. Ed., 2007, 46, 7860-7863. Copyright 2007 Wiley-VCH.]...

Powers ME, MatsuuraJ, Brassell J, Manning MC, and Shelter E. Enhanced Solubility of Proteins and Peptides in Nonpolar Solvents through Hydrophobic Ion Pairing. Biopolymers 1993 33 927-932. 

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