Chiral Recognition of Binaphthyl Derivatives with L-Undecyl Leucine Surfactants in the Presence of Sodium and Lysine Counterions

This study investigates the effect of counterions on the chiral recognition of 1,1'-Binaphthyl-2,2'-diamine (BNA) and 1,1'-Binaphthyl-2,2'-diyl hydrogenphosphate (BNP) enantiomers when using an amino acid-based surfactant undecanoyl L-leucine (und-Leu) as the chiral pseudostationary phase in capillary electrophoresis. The effects of using two different counterions (sodium and lysine) on the chiral recognition of binaphthyl derivatives were compared at varying pH conditions. The enantiomeric separation of BNA and BNP enantiomers via capillary electrophoresis, using und-Leu as the chiral recognition medium, significantly improved the enantiomeric resolution in capillary electrophoresis at pH 7 when using Lysine counterions as compared to using sodium as the counterion. More specifically, at a surfactant concentration of 45 mM, at pH 7, a significant increase in chiral selectivity was observed when lysine was used as the counterion compared to sodium. The enantiomeric resolution of BNA and BNP increased by 6-fold and 1.1-fold, respectively, in capillary electrophoresis experiments when lysine was utilized as the counterion compared to using sodium. Furthermore, the retention factor of BNA and BNP enantiomers also increased approximately 3.5-fold and 4-fold, respectively, in the presence of lysine counterions as compared to using sodium counterions. When running buffer in capillary electrophoresis was increased to pH 11, the resolution and retention factors were nearly identical when comparing the effects of the sodium and lysine counterions. This signifies the important role of lysine’s positive net charge on chiral recognition. This study provides insight into the potential advantages of using cationic, pH-dependent counterions such as lysine to significantly improve the chiral recognition of binaphthyl derivatives when using chiral anionic surfactants as the pseudostationary phase in capillary electrophoresis. How to cite this paper: Garcia, M., Risley, A., Billiot, F., Billiot, E. and Morris, K. (2021) Chiral Recognition of Binaphthyl Derivatives with L-Undecyl Leucine Surfactants in the Presence of Sodium and Lysine Counterions. American Journal of Analytical Chemistry, 12, 188-201. https://doi.org/10.4236/ajac.2021.125012 Received: February 24, 2021 Accepted: May 28, 2021 Published: May 31, 2021 Copyright © 2021 by author(s) and Scientific Research Publishing Inc. This work is licensed under the Creative Commons Attribution International License (CC BY 4.0). http://creativecommons.org/licenses/by/4.0/


Introduction
Chirality is ubiquitous in nature. Two of the most important simple class of chiral compounds include amino acids and sugars. Sugars and amino acids serve as the building blocks for a large percentage of biological compounds [1]- [6]. This suggests the profound influence of chirality on fundamental physiological and biological processes. Furthermore, stereospecific reactions play an essential role in drug metabolism, cell membrane stability, and gene expression [7]- [12].
Due to the aforementioned stereospecific reactions that occur within the body, it is not surprising that more than 50% of marketed drugs are chiral [13] [14] [15] [16]. These synthesized pharmaceutical drugs, however, frequently yield racemic mixtures, meaning it contains both enantiomers. This is of major concern because each enantiomer of a drug may exhibit different pharmacological effects. For example, the drug Thalidomide was originally marketed as a racemic mixture and administered to women to help mitigate the effects of morning sickness during pregnancy. Relatively soon after bringing this drug onto the market, an alarming rise in birth defects in newborns was observed. It was later determined that while one enantiomer caused the beneficial pharmaceutical effects, the other enantiomer was the cause of the teratogenic effects [17] [18]. In response to this tragedy, the Food and Drug Administration has mandated that each enantiomer of a chiral drug be evaluated for its respective physiological effects prior to being marketed [13] [16] [19] [20] [21] [22] [23]. This gave rise to a new scientific challenge: to establish and optimize enantiomeric separation processes and techniques.
Since then, many techniques have been established to separate enantiomeric compounds. Two of the more common techniques are high-performance liquid chromatography (HPLC) and capillary electrophoresis (CE) [19] [21] [24] [25] [26] [27]. CE typically yields a higher number of theoretical plates compared to HPLC and thus CE often yields better enantiomeric separations in a shorter period of time [28] [29]. In addition, CE allows for a quick and easy way to change the chiral recognition medium since the chiral recognition medium is part of the mobile phase acting as a pseudo-stationary phase. This is in contrast to HLPC which requires the purchase and installation of different analytical columns if one wishes to change the chiral recognition medium. With HPLC, a wide variety of chiral recognition media exist including but not limited to cyclodextrins, crown ethers, and chiral micelles [21] [34]. Amino acid-based surfactants are composed of a non-polar hydrocarbon chain and an amino acid head group [35]- [40]. The charge on the amino acid head groups can be greatly influenced by pH levels. The ability to impose different pH environments aids in studying how the charge of the surfactant, analytes, and counterions may affect the physiochemical properties of the micelles, as well as its ability to act as an effective chiral separation medium [30] [34]. All of which greatly influence the chiral recognition ability of the AABMs.
Previously, the effects of amino acid order, steric hindrance, dihedral angles, and the polymerization of AABMs have been studied experimentally and computationally to investigate their contributions to chiral recognition [ [34]. As previously reported from our research group, pH-dependent counterions, such as arginine have been demonstrated to significantly improve the enantiomeric resolution of various binaphthyl derivatives, when compared to using monatomic sodium as the counterion [34].
To further investigate the effects of pH-dependent counterions on chiral selectivity, this study investigates the role of lysine as the counterion on the chiral separations of 1,1'-Binaphthyl-2,2'-diyl hydrogenphosphate (BNP) and 1,1'-Binaphthyl-2,2'-diamine (BNA) enantiomers at pH 7 and 11 at varying concentrations of und-Leu surfactant. The structures of the surfactant, analytes, and lysine are provided in Figure 1. As shown in this figure, lysine contains two amine groups; one connected to the chiral carbon and another as a side chain substituent, which contains pKa values of approximately 9 and 10.5, respectively. As shown in Figure 2, lysine has various charge states at different pH levels. The various charge states can have a significantly different effect on the physicochemical properties and chiral recognition ability of the micelles formed. However, due to the solubility limitations of the micelles, we cannot work below pH 7, nor above pH 11 due to the limitations of capillary electrophoresis. Therefore, we limited our experiments to the useful pH extremes of our system, pH 7 and 11. Analyte standards were prepared in 1:1 methanol-water at 0.1 mg/mL. Samples were injected for 5 s at 10 mbar pressure. Separations were performed at +30 kV, with UV detection at 230 nm.

Results and Discussion
In this study, the chiral recognition of und-Leu at varying concentrations in the     Figure   5(a), no separation was achieved with sodium as the counterion but as shown in [34]. At pH 7, ~37% of the lysine molecules are bound to the surfactant. This number decreases to ~3% at pH 11 [30]. As seen in Figure 2, lysine has a net positive charge at pH 7, allowing for stronger electrostatic interactions between lysine and the negatively charged surfactants. However, this attraction is significantly reduced as it is subjected to higher pH levels. American Journal of Analytical Chemistry

Separation of BNP Enantiomers
The separation of the BNP enantiomers via MEKC with und-Leu surfactants in the presence of lysine and sodium counterions were compared at pH 7 and 11.
The resolution and k' values of the BNP enantiomers are shown in Figure 3 Figure 6. Contrarily, at pH 11, lysine has an overall net charge of zero thus losing its charged properties to act as an effective counterion. Due to the loss of positive charges, we therefore expect it to have less electrostatic and hydrogen bonding interactions with the negativelycharged surfactants. This is due to lysine losing its effectiveness as a counterion Figure 6. At pH 7, Lysine has a net charge of +2, therefore it is able to act as a counterion, bridging anionic surfactant head groups. Intermolecular bonding is not evident with sodium as it is monatomic.
at higher pH levels, which causes sodium ions to then act as the predominant counterion. Moreover, the interaction of BNP enantiomers with und-Leu is similar in the presence of sodium or neutralized lysine. Further evidence of similar interactions is shown in Figure 3(b), where similar retention factors for BNP were observed when comparing the presence of sodium or lysine counterions.

Separation of BNA Enantiomers
Similar to BNP, the BNA enantiomers exhibited better separation in the presence of lysine at pH 7. At pH 7, resolutions of 1.8, 2.0, 2.4 and 2.5 were observed at 15, 25, 35 and 45 mM, respectively, in the presence of lysine counterions, as presented in Figure 4(a). These resolution values far exceed those observed in the presence of sodium at the same pH level. As seen in Figure 4(a), the BNA enantiomers were separated with resolutions of 0.4, 0.9, 1.6 and 2.1 at concentrations of 15, 25, 35 and 45 mM, respectively in the presence of sodium counterions. As previously mentioned, the CMC of und-Leu is approximately the same (~17 -18 mM) in the presence of sodium and lysine. However, at pH 7, baseline separation of BNA enantiomers was observed at 15 mM. Furthermore, the k' values indicate that BNA enantiomers do not bind strongly to und-Leu at this surfactant concentration. Overall, the interaction of BNA enantiomers is very effective at low pH levels and low surfactant concentrations. Below the CMC, lysine counterions still interact with und-Leu surfactants. At pH 7, lysine molecules are positively charged, therefore, electrostatic attraction between the positively charged amine moieties of lysine and the negatively charged und-Leu draw these two molecules closer together. This allows lysine to form hydrogen bonds with the surfactant polar head groups thus providing a chiral cavity that can improve its chiral selectivity of BNA enantiomers.

Effect of Lysine Chirality on Chiral Recognition
To determine if the chirality of lysine played a role in chiral recognition, two experiments were performed. In one experiment, 50 mM L-Lysine was used without und-Leu, and no chiral recognition was observed. In another experi-American Journal of Analytical Chemistry

Conclusions
The enantiomeric resolution of BNP significantly improved in the presence of lysine counterions as compared to sodium counterions at pH 7 when using und-Leu surfactants as the chiral recognition medium. Most notably, when using a surfactant concentration of 45 mM in the presence of lysine counterions at pH 7, the enantiomeric resolution increased approximately 6-fold compared to that of when using sodium as the counterion. Furthermore, at the same conditions, the retention factor increased approximately 4-fold when using lysine counterions, as compared to using sodium. However, with experimental conditions at pH 11, the enantiomeric resolution and retention factor are nearly identical when separating BNP enantiomers with either lysine or sodium counterions. At pH 7, the enantiomeric resolution of BNA enantiomers was achieved using und-Leu surfactants in the presence of lysine counterions. This improved the enantiomeric resolution values as compared to when using sodium counterions. For example, at a surfactant concentration of 45 mM in the presence of lysine at pH 7, the enantiomeric resolution increased approximately 1.1-fold compared to that of when using sodium counterions. Furthermore, the retention factor at those same aforementioned conditions increased approximately 3.5-fold when using lysine as the counterion, as compared to using sodium. When separating BNA enantiomers using 45 mM concentration of und-Leu surfactant at pH 11, the enantiomeric resolution and retention factors are nearly identical to that of when using sodium as the counterion.
In conclusion, when separating chiral compounds such as BNA and BNP with und-Leu surfactants in the presence of pH-dependent counterions such as Lysine via MEKC, the enantiomeric resolution and retention factors significantly improved when compared to using sodium counterion as the counterion in these studies. Therefore, this study provides insight to further optimize chiral separation conditions using pH-dependent counterions as opposed to monatomic counterions such as sodium.