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Establishment and molecular modeling study of maltodextrin-based synergistic enantioseparation systems with two new hydroxy acid chiral ionic liquids as additives in capillary electrophoresis
Accepted Manuscript Title: Establishment and molecular modeling study of Maltodextrin-based synergistic enantioseparation systems with two new hydroxy acid chiral ionic liquids as additives in capillary electrophoresis Authors: Xuan Yang, Yingxiang Du, Zijie Feng, Zongran Liu, Jingtang Li PII: DOI: Reference:
S0021-9673(17)30840-3 http://dx.doi.org/doi:10.1016/j.chroma.2017.06.007 CHROMA 358577
To appear in:
Journal of Chromatography A
Received date: Revised date: Accepted date:
8-3-2017 26-5-2017 1-6-2017
Please cite this article as: Xuan Yang, Yingxiang Du, Zijie Feng, Zongran Liu, Jingtang Li, Establishment and molecular modeling study of Maltodextrinbased synergistic enantioseparation systems with two new hydroxy acid chiral ionic liquids as additives in capillary electrophoresis, Journal of Chromatography Ahttp://dx.doi.org/10.1016/j.chroma.2017.06.007 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Establishment and molecular modeling study of Maltodextrin-based synergistic enantioseparation systems with two new hydroxy acid chiral ionic liquids as additives in capillary electrophoresis Xuan Yang1, Yingxiang Du1, 2, 3, *, Zijie Feng1, Zongran Liu1, Jingtang Li1
1
Department of Analytical Chemistry, China Pharmaceutical University, Nanjing
210009, P. R. China 2
Key Laboratory of Drug Quality Control and Pharmacovigilance (Ministry of
Education), China Pharmaceutical University, Nanjing 210009, P. R. China 3
State Key Laboratory of Natural Medicines, China Pharmaceutical University,
Nanjing 210009, P. R. China
*
Correspondence: Professor Yingxiang Du, Department of Analytical Chemistry,
China Pharmaceutical University, No.24 Tongjiaxiang, Nanjing, Jiangsu 210009, China E-mail: [email protected] Tel./fax: +86 25 83221790
Highlights Two new chiral hydroxy acid-based ionic liquids (tertramethylammonium -D-pantothenate and tertramethylammonium-D-quinate) were designed and first used as additives to construct the maltodextrin-based synergistic system for enantioseparation in capillary electrophoresis (CE).
Compared to traditional system, significantly improved separations in the CIL/Maltodextrin synergistic systems were obtained.
The chiral recognition process of the synergistic systems based on maltodextrin was first evaluated by molecular modeling method.
The modeling results were in good agreement with our experimental results.
Abstract： Discovering more superior performance of ionic liquids for the separation science has triggered increasing interest. In this work, two new Hydroxy
acid-based
chiral
ionic
liquids
(tertramethylammonium-D-pantothenate
(CILs)
(TMA-D-PAN),
tertramethylammonium-D-quinate (TMA-D-QUI)) were designed and first used as additives to establish the maltodextrin-based synergistic systems for enantioseparation in capillary electrophoresis (CE). Compared to traditional single maltodextrin chiral separation system, significantly improved separations of
all tested drugs in the
CIL/Maltodextrin synergistic systems were obtained. Some parameters (CIL concentration, maltodextrin concentration, buffer pH, and applied voltage) in the TMA-D-PAN/Maltodextrin synergistic system have been examined and optimized for analytes. The molecular docking software AutoDock was applied to simulate the recognition process and surmise feasible resolution mechanism in the Maltodextrin/CILs synergistic systems, which has certain guiding value.
Keyword:
Chiral
ionic
liquid;
Synergistic
system;
electrophoresis; Enantioseparation; Molecular docking
Capillary
1. Introduction Chirality is one of the basic characteristics of nature. Almost half of the drugs in current use are known to be chiral and only a small number are administered as pure enantiomers. The popularity of the research on single enantiotopic chiral drugs stems from the fact that the desired pharmacological activity is exhibited by one enantiomer, while the other enantiomer may exhibit low activity, opposite pharmacokinetic and pharmacodynamics effects, or even toxicity. Among the various analytical techniques for enantioseparation, capillary electrophoresis (CE) has become a powerful alternative to chromatography, owing to its several advantages such as high resolving power, rapid analysis, low consumption of sample, solvent, and chiral selector, as well as high flexibility in choosing and changing types of selectors and separation modes[1]. The most common approaches for enantiomeric separation in CE involve the addition of one or more chiral selectors into the running buffer. Until now, various kinds of chiral selectors, including cyclodextrins and their derivatives[2-5], chiral crown ethers[6, 7], polysaccharides[8-10], antibiotics [11-13] and many other small molecules have been explored in CE for enantioseparation. However, in some cases, satisfactory results may not been obtained in the conventional separation systems using single chiral selector. Accordingly,
a number of research groups focus on hunting for different types of additives to enhance their enantioselectivity. Polysaccharides always possess variable structures and numerous functional chiral groups. Additionally, they have properties of the water-soluble and weak UV absorption, which benefit the application of these materials as CE chiral additives for enantioseparation.[9,10]. Maltodextrin, as an optically active polysaccharide chiral selector, can form the helical structure with hydrophilic outside and hydrophobic cavity in acidic and neutral aqueous solution[14].Compared with the rigid cavity of cyclodextrins, the spiral structure of maltodextrin, varying along with the size of the guest molecules, could be more flexible in chiral recognition[15,16].
However,
the
disadvantages
of
the
single
maltodextrin system (like high concentration, and finite enantioseparation capability) limit its application. Room-temperature ionic liquids (ILs), which are generally defined as organic salts in the liquid state under the condition of close to or below room temperature[17], have been developed fast in separation science, such as stationary phase in gas chromatography[18], mobile phase additives
in
liquid
chromatography[19],
solvent
in
two-phase
extraction[20]. In CE, ILs have been used as electrolyte[21, 22], running buffer modifiers[23], surface coating or chemical modification of the
capillary wall[24]. Chiral ionic liquids (CILs) are a subclass of ILs that have a either chiral cation, chiral anion, or both. On account of their potential chiral discrimination capabilities, the scientific interest is continuously increasing. However, in the recent years, only a few papers have reported the application of chiral ILs to establishment synergistic systems for enantioseparation in CE [25-30], and most of them are amino acid ionic liquids (AAILs) or their derivatives [26, 27]. As far as we know, only two papers about hydroxy acid ionic liquid as additives of the synergistic system [28, 29] were reported. In recent years, some studies have provided good insights into the host-guest interactions between CDs and the enantiomers and the chiral recognition mechanisms with molecular modeling methods[31-33], nuclear magnetic resonance (NMR)[34], X-ray crystallography[35]. Molecular modeling methods which included molecular dynamics simulations, Parametric Model 3(or 6) semiempirical methods, and molecular docking, have been performed to rationalize experimental findings. Moreover, molecular modeling methods have been successfully applied to evaluate the influence of hydrogen bonding in the chiral recognization process, which is difficult to explain because of the limitation of experimental methods. Investigations have never been reported which apply molecular modeling methods to elucidate the
possible chiral recognition mechanism of the polysaccharide-based synergistic systems. In
this
paper,
two
newly
tetramethylammonium-D-pantothenate tetramethylammonium-D-quinate
designed
hydroxy
acid
CILs
(TMA-D-PAN)
(TMA-D-QUI)
were
applied
and as
additives to evaluate their potential synergistic effect with maltodextrin as the chiral selector in CE. For the further understanding of the possible chiral recognition mechanism of synergistic system, we examined the course of host-guest inclusion between maltodextrin and enantiomers by means of a molecular docking technique and calculated the binding free energy by the AutoDock semi-empirical binding free energy function. The modeling results were in good agreement with our experimental results. 2. Experimental 2.1 Chemicals and reagents Maltodextrin (M040，DE4-7) was purchased from Sigma (St. Louis, Mo,
USA).Tetramethylammonium-D-pantothenate
Tetramethylammonium-D-quinate
(TMA-D-PAN),
(TMA-D-QUI)
and
Tetramethylammonium hydroxide (TMA-OH) ionic liquid (purity > 98%) were purchased from Shanghai Cheng Jie Chemical Co., Ltd. (Shanghai, China). Nefopan(NEF), Ketoconazole (KET), Econazole (ECO) and
Voriconazole(VOR), were supplied by Jiangsu Institute for Food and Drug Control (Nanjing, China). All these drug samples were racemic mixtures. The chemical structures of those substances are shown in Fig.1. Tris (hydroxymethyl) aminomethane (Tris) were purchased from Shanghai Huixing Biochemistry Reagent (Shanghai, China). Phosphoric acid and sodium hydroxide were of analytical regent from Nanjing Chemical Reagent (Nanjing, China). Double distilled water was used throughout all the experiments. Figure 1 2.2. Apparatus Electrophoretic experiments were performed with an Agilent 3D CE system (Agilent Technologies, Waldbronn, Germany), which consisted of a sampling device, a power supply, a photodiode array UV detector (wavelength range from 190 nm to 600 nm) and a date processor. The whole system was driven by Agilent ChemStation software (Revision B.02.01) for system control, date collection and analysis. It was equipped with a 50 cm (41.5 cm effective length) × 50 μm id uncoated fused-silica capillary (HebeiYongnian County Reafine Chromatography Ltd., Hebei, China). The samples were introduced hydrodynamically for 2 s (injection pressure 50 mbar). All separations were carried out at 20 oC using a voltage in the range of 20-28 kV. The wavelength for detection was 220
nm (NEF), 215 nm (KET, ECO, and VOR). The CE system was operated in the conventional mode with the anode at the injector end of the capillary. A new capillary was first rinsed with 1.0 M NaOH (30 min), followed by 0.1 M NaOH (20 min) and water (20 min), respectively. At the beginning of each day, the capillary was flushed with 0.1 M NaOH (10 min) followed by water (10 min). Between consecutive injections, the capillary was rinsed with 1.0 M NaOH, 0.1 M NaOH, water and running buffer for 3 min each. Nylon filters (0.45 μm) were purchased from Jiangsu Hanbon Science and Technology (Nanjing, China). All the solution need to be filtered before used. 2.3 Procedures The background electrolyte (BGE) consisted of 60 mM Tris solution (if not stated otherwise), adjusted to specified pH value with H 3PO4 (20%, v/v). The running buffer solutions were freshly prepared by dissolving appropriate amounts of maltodextrin and/or ILs in BGE, and then adjusting pH exactly to a desired value by adding a small volume of H3PO4 (20%, v/v) using a microsyringe. The racemic sample of NEF was dissolved
in
distilled
water.
The
racemic
samples
of
azole
antifungals(KET, ECO,VOR) were dissolved in methanol (50%，v/v).All those racemic samples were prepared at a concentration of 1.0 mg/ml.
Running buffers and samples were filtered with a 0.45 μm pore membrane filter and degassed by sonication prior to use. 2.4 Molecular Construction and Optimization Molecular construction was carried out using the ChemOffice 2010 software package. The MM2 force field and molecular dynamics were applied for the structure optimization. The PM3 quantum mechanical method was used for geometry optimization to adjust any spatially unreasonable bond distances and bond angles of all enantiomers. Glucose-α (1→4)-glucose is the backbone glycosidic linkage in maltodextrin [15,16]. The φ and ψ dihedral angles for the α(1→4)-linkage are define as φ = H1—C1—O1—C4＇, φ = C1—O1—C4＇—H4＇.The number of D-glucose units in the three-dimensional structure of the maltodextrin was set at 15 (based on the average of molecular weight)and the
crystal
structure
values
of
φ,ψ=5°,13°
in
the
vacuum
condition[36-38].Geometry optimization was conducted by applying the PM3 quantum mechanical method. 2.5 Molecular docking simulations Molecular docking simulations were performed with the automated docking program of AutoDock 4.2.3[39]. It employed the Lamacrkian genetic algorithm (LGA) to identify binding conformations of a flexible ligand (or small molecule) to a target receptor. The two CILs
(TMA-D-PAN and TMA-D-QUI)studied in this work were supposed to exist in two states of ionic associated and dissociated, resulting from the fact that ionic liquids are partially dissociated in the aqueous solutions due to the dual nature of ILs (hydrophilic/hydrophobic properties) associated to non-polar and ionic interactions[40].To comprehensive investigate the effect of the two chiral ionic liquids，associated CILs and dissociated CILs were separately introduced in the molecular model construction. As a preliminary preparation step to docking, AutoGrid[41] creates 3D grid boxes to generate a simplified representation for target receptor (e.g. maltodextrin). Each atom type (called probe) of the ligand will be placed at the grid points and its interaction energy with all the atoms of the receptor will be computed and assigned to the corresponding grid point[42]. Grid maps of dimensions70 A˚×60 A˚×60 A˚, with a grid spacing of 0.375 A˚, were placed to cover the maltodextrin molecule. One hundred LGA runs, each with 200 individuals in the population, were performed. Results differing by less than 1 A˚ in a positional root mean square deviation (rmsd) were clustered together. In each group, the lowest binding energy configuration with the highest % frequency was selected as the group representative. 3. Results and discussion 3.1 Effect of different chiral ILs and achiral IL
The electropherograms of four enantiomers (NEF, KET, ECO, and VOR) in different chiral separation systems was shown in Fig.2. Under the optimal conditions (60 mM Tris-H3PO4 buffer solution containing 3.0% (w/v) maltodextrin, 50 mM TMA-D-PAN or 50 mM TMA-D-QUI ; buffer pH 3.0; applied voltage, 24 kV for NEF, 26 kV for KET, ECO, VOR; capillary temperature, 20 °C), the entirely separations of all the tested drugs were obtained in the synergistic systems. As observed in Table 1, with the EOF reversed, the migration time of the analytes in ILs/Maltodextrin systems were longer than counterparts in the single maltodextrin system. This results from the increased ionic liquid strength of the running buffer, as well as the adsorption of IL cations (TMA+) on the capillary inner surface [43, 44]. The prolong migration time would provide more opportunities for chiral recognition in the separation process. A comparative study was conducted between achiral IL (TMA-OH)/Maltodextrin system and chiral ILs/Maltodextrin system. As observed, the addition of TMA-OH can also reverse the EOF in a similar extent. But the TMA-OH/Maltodextrin synergistic system exhibited poorer enantioseparation than the CILs/Maltodextrin systems, indicating the indispensability of the chiral parts of CILs in the synergistic systems. As excepted, significantly improved Rs and α of all the tested enantiomers were obtained in both of the TMA-D-PAN/Maltodextrin
system and TMA-D-QUI/Maltodextrin system, suggesting the existence of the synergistic effect between CILs and chiral selector maltodextrin. With regard to the two studied CILs, TMA-D-PAN exhibited better synergistic effect towards four analytes than TMA-D-QUI. It might be attributed to the chiral anion part of CILs, which is the only disparity between the structures of two CILs. This phenomenon indicated that the structure and properities of the chiral part of chiral ILs have a significant impact on the chiral discrimination process of the CILs synergistic systems [26-30]. It is worth noting that no enantioseparation for all tested drug were obtained with only CILs. To test the repeatability of the chiral separation systems, including single Maltodextrin, achiral IL/Maltodextrin, and CILs/Maltodextrin synergistic separation systems, five consecutive separations of four enantiomers were performed. For all studied analytes, the intra- and inter day variations of Rs (the RSD values of the Rs) were less than 3.6% and 3.9%, respectively. The intra- and interday variations of migration times (the RSD values of the migration times) were less than 3.3% and 3.7%， respectively. It could therefore be concluded that the synergistic systems could provide satisfactory repeatability. Table 1 Figure 2
3.2
Effect
of
CIL
concentration
on
enantioseparation
TMA-D-PAN/Maltodextrin synergistic system was selected to optimize the separation conditions. To investigate the effect of CIL concentration on enantioseparation, experiments were carried out by varying the CIL concentration in the range of 20 mM-60 mM while keeping maltodextrin concentration constant at 3.0% （w/v）. As observed in Table 2, the migration time of all the enantiomers increased with the CIL concentration changing from 20 mM to 60 mM mainly owing to a decrease in the density of negative charge resulting from the adsorption of the IL cations on the surface of the fused-silica capillary and, especially, the increased complexation (synergistic effect) between enantiomers and chiral recognition reagents. With the increase of migration time, the separation resolution and selectivity of NEF, KET, ECO, VOR, increased gradually with CIL concentration rising from 20 mM to 50 mM, indicating that the synergistic effect between chiral IL and maltodextrin continuously improved. At higher CIL concentration (50 mM-60 mM), the selectivity of KET, ECO decreased, that observation probably was owned to the saturation of the synergistic effect between CIL and maltodextrin in the concentration of 50 mM. While the selectivity of NEF, VOR continued to increase in 60 mM, the separation
resolution declined presumably, due to the minute broadening peak at high CILs concentration. Taking account of good separation and appropriate migration time simultaneously, the CIL concentration of 50 mM was determined to optimum for the synergistic system. Table 2 3.3 Effect of maltodextrin concentration on enantioseparation The concentration of chiral selectors is also an important factor influencing the enantioseparation. To obtain the optimal selector concentration of the TMA-D-pantothenate/Maltodextrin synergistic system, we investigated a series of concentrations (1.5%-3.5% (w/v)) of Maltodextrin with 60 mM Tris/H3PO4 buffer solution (pH 3.0) containing 50 mM TMA-D-pantothenate. At the same time, we studied the single maltodextrin system, and all studied drugs were partially separated. As shown in Fig.3, the Rs and α of all the racemic drugs initially increased along with concentration of chiral selector increasing from 1.5% (w/v) to 3.0% (w/v), which was probably imputed to the change of solution viscosity and the enhancement of interaction among maltodextrin, CIL and enantiomers. Then the Rs and α of three chiral pharmaceutical (NEF, KET, ECO) decreased at high concentration (3.5% (w/v)), indicating the oversaturated situation of the synergistic system. The selectivity of VOR
continuously increased, while the resolution declined mainly due to the fact that it appeared to sustain the peaking broadening. Moreover, compared with the single maltodextrin system, TMA-A-PAN/Maltodextrin synergistic system could achieve better separation resolution under all the experimental conditions of maltodextrin concentration. By the results, 3.0% (w/v) maltodextrin was selected for further analysis. Figure 3 3.4 Effect of buffer pH on enantioseparation Buffer
pH
is
a
critical
and
sensitive
parameter
for
the
CIL/Maltodextrin synergistic system optimization due to the significant influence in the ionic state of capillary wall, the dissociation of analytes, the electrophoretic behavior of solutes and the interaction among chiral selectors, chiral ILs and enantiomers (e.g. electrostatic interaction and hydrogen bondings). The effect of the BGE pH on chiral separation was investigated by increasing the pH from 2.0 to 4.0 using 60 mM Tris/H3PO4 containing 3.0% (w/v) maltodextrin and 50 mM TMA-Dpantothenate, while all the analytes with a positive charge. As can be seen from Fig.4, the separation resolutions and selectivities for all the chiral drugs increased when pH increased from 2.0 to 3.0 indicating the enhancement of the synergistic system effect, and then they decreased
when pH further increased from 3.0 to 4.0. This observation could be explained by the fact that the enhancement of the synergistic system effect with the pH increased, thus chiral resolution and selectivity was increased. However, further increasing of pH would mainly weaken the interaction among chiral selectors, chiral ILs and enantiomers. Therefore, buffer pH at 3.0 was selected for further analysis. Figure 4 3.5 Effect of applied voltage on enantioseparation In CE, applied voltage is an important factor to control the column efficiency, the resolution, and the analysis time. Higher voltage would bring about higher CE efficiency and shorter migration time, while it leaded to encumbered Joule heating, which mainly reduced the resolution. In another way, the prolonged migration time along with lower applied voltage would give more chances for the enantiorecognition, which always influences the resolution. In order to evaluate the effect of applied voltage on chiral separation of the synergistic system, we studied a series of voltage (20 kV-28 kV) with 60 mM Tris/H3PO4 buffer solution (pH 3.0) containing 50mM TMA-D-pantothenate and 3% (w/v) maltodextrin. As shown in Fig.5, the resolutions of all racemic drugs initially increased owning to the improvement of CE efficiency with the applied voltage increasing, and
then the decline of resolution appeared probably because of extra Joule heating and peaking broadening. Considering enantioseparation and migration time, we selected applied voltage of 24 kV for NEF and 26 kV for KET, ECO, VOR in the synergistic system. Figure 5 4. Molecular modeling study of chiral recognition mechanism In this study, we used the molecular docking software AutoDock 4.2.3 to confirm and explain the possible chiral recognition mechanism and the potential synergistic effect of the CILs/Maltodextrin synergistic system. The molecular docking configurations for KET enantiomers in single maltodextrin synergistic
system system
(b)
(a), and
dissociated-TMA-D-PAN/Maltodextrin associated-TMA-D-PAN/Maltodextrin
synergistic system (c) are presented in Fig.6. Generally, there are multiple interactions participating in the process of the enantiorecognition, including hydrogen bonding interaction, dipole-dipole, π-π, charge-charge, host-guest inclusion, etc. Although not all the interactions could be observed directly, the molecular docking configurations of the inclusion complexes still provide visualization of some interactions for the investigation of chiral recognition mechanism. As shown in Fig.6, hydrogen bonding interactions (labeled by the green line) for KET were dramatically enhanced in TMA-D-PAN/Maltodextrin synergistic systems
(no matter what state of TMA-D-PAN is) compared with single maltodextrin system. Meanwhile, the σ-π binding interaction (indicated by orange line) appeared between enantiomer and maltodextrin in associated TMA-D-PAN/Maltodextrin synergistic system. The results above suggested the CILs truly have impact on the recognition of maltodextrin. The molecular docking results with AutoDock for all tested enantiomers in both single maltodextrin and CILs/Maltodextrin synergistic systems are presented in Table 3. Thermodynamically, the more negative of binding energies indicates the more stable state of the inclusions. As shown in Table 3, whatever the state of CILs is, the addition of CILs to the single maltodextrin system improved the values of |∆G| for all studied drugs. Moreover, the results showed that the participation of dissociated and associated CILs could increase the difference of binding free energies (|∆∆G|) between enantiomers significantly, which might be attributed to the increased chiral discrimination and improved chiral selectivity in maltodextrin separation systems. Simultaneously, the difference of the calculated binding energy corresponded with the experimental chiral selectivity α (see Table 1). In addition, the values of |∆∆G| in the associated CILs/Maltodextrin system were higher than those in the dissociated CILs/Maltodextrin system for
all four enantiomers, indicating the essential role of associated CILs in CILs/Maltodextrin synergistic system for chiral separation. Above all, the existence of the CILs may ameliorate the chiral environment, and thus improve the chiral recognition ability and selectivity of the original system, indicating the synergistic effect in the CILs/Maltodextrin. Figure 6 Table 3
5. Conclusions In this paper, two novel chiral ILs based on hydroxyl acid ， TMA-D-PAN and TMA-D-QUI, were first used as additives to establish the Maltodextrin-based synergistic systems in CE for enantioseparation. Some parameters (CIL concentration, maltodextrin concentration, buffer pH, and applied voltage) in the TMA-D-PAN/Maltodextrin synergistic system have been examined and optimized for all tested drugs. Significant improvements of chiral separation for analytes were obtained with CILs compared to the single maltodextrin system. The chiral recognition mechanism of the CILs/Maltodextrin synergistic systems were further investigated with the molecular docking software AutoDock 4.2.3., which serves as a good method for predicting enantioseparation.
Acknowledgements This work was supported by the Project of National Natural Science Foundation of China (No.: 81373378) and the Natural Science Foundation of Jiangsu Province (Program No.BK20150697) and the Project of the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD).
The authors have declared no conflict of interst
Figure Caption
Fig.1 Structures of (A) Maltodextrin (B) TMA-OH (C) TMA-D-PAN (CIL1) and TMA-D-QUI (CIL2) (D) Four racemic drugs.
Fig.2 Typical electrophoregrams of the chiral separation of (A) NEF, (B) KET, (C) ECO, (D) VOR in the different separation system. Conditions: 60 mM Tris-H3PO4 buffer solution containing (a) 3.0% (w/v) maltodextrin; (b) 3.0% (w/v) maltodextrin and 50 mM D-PAN-Na+; (c) 3.0% (w/v) maltodextrin and 50 mM TMA-OH; (d) 3.0% (w/v) maltodextrin and 50 mM TMA-D-PAN; (e) 3.0% (w/v) maltodextrin and 50 mM TMA-D-QUI; buffer pH 3.0; applied voltage, 24 kV (NEF), 26 kV (KET, ECO, VOR); capillary temperature, 20 oC other condition see section 2.
Fig.3 Effect of chiral selector concentration on the enantioseparation. Conditions: 60 mM Tris-H3PO4 buffer solution (pH 3.0) containing 1.5%-3.5% (w/v) maltodextrin with 50 mM CIL1 (TMA-D-PAN); applied voltage, 24 kV (NEF), 26 kV (KET, ECO, VOR); capillary temperature, 20 °C; other condition see section 2.
Fig.4 Effect of buffer pH on the enantioseparation. Conditions: 60 mM
Tris-H3PO4 buffer solution containing 3.0% (w/v) maltodextrin and 50 mM TMA-D-PAN; buffer pH 2.0-4.0; applied voltage, 24 kV (NEF), 26 kV (KET, ECO, VOR); capillary temperature, 20 °C; other condition see section 2.
Fig.5 Effect of applied voltage on the enantioseparation. Conditions: 60 mM Tris-H3PO4 buffer solution containing 3.0% (w/v) maltodextrin and 50 mM TMA-D-PAN; buffer pH 3.0; applied voltage, 20 kV-28 kV; capillary temperature, 20 °C; other condition see section 2.
Fig. 6 Molecular docking configuration for KET enantioseparation in a single Maltodextrin system; b dissocated TAM-D-PAN/Maltoextrin synergistic
system
and
c
associated
TMA-D-PAN/Maltodextrin
synergistic system. The left is (R)-enantiomer and the right is (S)-enantiomer. The hydrogen bonding is indicated by green dotted line. The σ-π is indicated by orange line. O red，N blue，H white，C grey.
References [1] Koppenhoefer B, Epperlein U, Christian B, et al. Separation of enatiomers of drugs by capillary electrophoresis III. β-cyclodextrin as chiral solvating agent [J]. Journal of Chromatography A, 1996, 735(1-2): 333-343. [2]Saz J M, Marina M L. Recent advances on the use of cyclodextrins in the chiral analysis of drugs by capillary electrophoresis [J]. Journal of Chromatography A, 2016, 1467: 79-94. [3] Zhu Q, Scriba G K E. Advances in the use of cyclodextrins as chiral selectors in capillary electrokinetic chromatography: fundamentals and applications [J]. Chromatographia, 2016, 79(21-22): 1403-1435. [4] Řezanka P, Navrátilová K, Řezanka M, et al. Application of cyclodextrins in chiral capillary electrophoresis [J]. Electrophoresis, 2014, 35(19): 2701-2721. [5] Escuder-Gilabert L, Martín-Biosca Y, Medina-Hernández M J, et al. Cyclodextrins in capillary electrophoresis: recent developments and new trends [J]. Journal of Chromatography A, 2014, 1357: 2-23. [6]Paik M J, Kang J S, Huang B S, et al. Development and application of chiral crown ethers as selectors for chiral separation in high-performance liquid chromatography and nuclear magnetic resonance spectroscopy [J]. Journal of Chromatography A, 2013, 1274: 1-5. [7] Kuhn R. Enantiomeric separation by capillary electrophoresis using a
crown ether as chiral selector [J]. Electrophoresis, 1999, 20(13): 2605-2613. [8] Nishi H, Izumoto S, Nakamura K, et al. Dextran and dextrin as chiral selectors in capillary zone electrophoresis [J]. Chromatographia, 1996, 42(11): 617-630. [9] Chen J, Du Y, Zhu F, et al. Glycogen: a novel branched polysaccharide chiral selector in CE [J]. Electrophoresis, 2010, 31(6): 1044-1050. [10] Zhang Q, Du Y, Chen J, et al. Investigation of chondroitin sulfate D and chondroitin sulfate E as novel chiral selectors in capillary electrophoresis [J]. Analytical and bioanalytical chemistry, 2014, 406(5): 1557-1566. [11]Chen B, Du Y, Wang H. Study on enantiomeric separation of basic drugs by NACE in methanol ‐ based medium using erythromycin lactobionate as a chiral selector[J]. Electrophoresis, 2010, 31(2): 371-377. [12] Domínguez‐Vega E, Pérez‐Fernández V, Crego A L, et al. Recent advances in CE analysis of antibiotics and its use as chiral selectors [J]. Electrophoresis, 2014, 35(1): 28-49. [13]Domínguez‐Vega E, Montealegre C, Marina M L. Analysis of antibiotics by CE and their use as chiral selectors: An update [J]. Electrophoresis, 2016, 37(1): 189-211. [14] Nishi H, Izumoto S, Nakamura K, et al. Dextran and dextrin as chiral
selectors in capillary zone electrophoresis[J]. Chromatographia, 1996, 42(11): 617-630. [15] Stefan R I, Van Staden J F, Aboul-Enein H Y, et al. Maltodextrins as new chiral selectors in the design of potentiometric, enantioselective membrane electrodes[J]. Fresenius' journal of analytical chemistry, 2001, 370(1): 33-37. [16] Tabani H, Fakhari A R, Nojavan S. Maltodextrins as chiral selectors in CE: Molecular structure effect of basic chiral compounds on the enantioseparation[J]. Chirality, 2014, 26(10): 620-628. [17] Tang S, Liu S, Guo Y, et al. Recent advances of ionic liquids and polymeric ionic liquids in capillary electrophoresis and capillary electrochromatography[J]. Journal of Chromatography A, 2014, 1357: 147-157. [18] Zhang C, Park R A, Anderson J L. Crosslinked structurally-tuned polymeric ionic liquids as stationary phases for the analysis of hydrocarbons
in
kerosene
and
diesel
fuels
by comprehensive
two-dimensional gas chromatography [J]. Journal of Chromatography A, 2016, 1440: 160-171. [19] Wang Q, Chen X, Qiu B, et al. Ionic liquid as a mobile phase additive in high ‐ performance liquid chromatography for the simultaneous determination of eleven fluorescent whitening agents in paper materials[J]. Journal of Separation Science, 2016, 39(7):1242.
[20]Tang F, Zhang Q, Ren D, et al. Functional amino acid ionic liquids as solvent and selector in chiral extraction [J]. Journal of Chromatography A, 2010, 1217(28): 4669-4674. [21] Zhang H, Qi L, Mu X, et al. Influence of ionic liquids as electrolyte additives on chiral separation of dansylated amino acids by using Zn (II) complex mediated chiral ligand exchange CE[J]. Journal of separation science, 2013, 36(5): 886-891. [22] Liu R, Du Y, Chen J, et al. Investigation of the enantioselectivity of tetramethylammonium L-hydroxyproline ionic liquid as a novel chiral ligand in ligand-exchange CE and ligand-exchange MEKC[J]. Chirality, 2015, 27(1): 58-63. [23]Borissova M, Palk K, Koel M. Micellar electrophoresis using ionic liquids [J]. Journal of Chromatography A, 2008, 1183(1): 192-195. [24]Jin Y, Chen C, Meng L, et al. Simultaneous and sensitive capillary electrophoretic
enantioseparation
of
three
β-blockers
with
the
combination of achiral ionic liquid and dual CD derivatives[J]. Talanta, 2012, 89: 149-154. [25] François Y, Varenne A, Juillerat E, et al. Evaluation of chiral ionic liquids as additives to cyclodextrins for enantiomeric separations by capillary electrophoresis[J]. Journal of Chromatography A, 2007, 1155(2): 134-141. [26] Zhang J, Du Y, Zhang Q, et al. Investigation of the synergistic effect
with amino acid-derived chiral ionic liquids as additives for enantiomeric separation in capillary electrophoresis [J]. Journal of Chromatography A, 2013, 1316: 119-126. [27] Zhang Q, Du Y. Evaluation of the enantioselectivity of glycogen-based synergistic system with amino acid chiral ionic liquids as additives in capillary electrophoresis [J]. Journal of Chromatography A, 2013, 1306: 97-103. [28] Zuo L, Meng H, Wu J, et al. Combined use of ionic liquid and β‐ CD for enantioseparation of 12 pharmaceuticals using CE [J]. Journal of separation science, 2013, 36(3): 517-523 [29] Cui Y, Ma X, Zhao M, et al. Combined Use of Ionic Liquid and Hydroxypropyl ‐ β ‐ Cyclodextrin for the Enantioseparation of Ten Drugs by Capillary Electrophoresis [J]. Chirality, 2013, 25(7): 409-414. [30] Zhang Y, Du S, Feng Z, et al. Evaluation of synergistic enantioseparation systems with chiral spirocyclic ionic liquids as additives by capillary electrophoresis[J]. Analytical and bioanalytical chemistry, 2016, 408(10): 2543-2555. [31] Li W, Liu C, Tan G, Zhang X, Zhu Z, Chai Y, Molecular modeling study of chiral separation and recognition mechanism of beta-adrenergic antagonists by capillary electrophoresis, Int. J. Mol. Sci, 13 (2012) 710-725. [32] Alvira E, Molecular dynamics study of the influence of solvents on
the chiral discrimination of alanine enantiomers by β-cyclodextrin, Tetrahedron: Asymmetry, 24 (2013) 1198-1206. [33] Nascimento C S, Lopes J F, Guimarães L, et al. Molecular modeling study of the recognition 4-hydroxypropranolol
mechanism and
by
capillary
enantioseparation of
electrophoresis
using
carboxymethyl-β-cyclodextrin as the chiral selector[J]. Analyst, 2014, 139(16): 3901-3910. [34] Li W, Tan G, Zhao L, et al. Computer-aided molecular modeling study of enantioseparation of iodiconazole and structurally related triadimenol analogues by capillary electrophoresis: chiral recognition mechanism and mathematical model for predicting chiral separation[J]. Analytica chimica acta, 2012, 718: 138-147. [35] Stiufiuc R, Iacovita C, Stiufiuc G, et al. Surface mediated chiral interactions between cyclodextrins and propranolol enantiomers: a SERS and DFT study[J]. Physical Chemistry Chemical Physics, 2015, 17(2): 1281-1289. [36] Quigley G J, Sarko A, Marchessault R H. Crystal and molecular structure of maltose monohydrate [J]. Journal of the American Chemical Society, 1970, 92(20): 5834-5839. [37] Kuttel M M, Naidoo K J. Free energy surfaces for the α (1→ 4)-glycosidic linkage: implications for polysaccharide solution structure and dynamics [J]. The Journal of Physical Chemistry B, 2005, 109(15):
7468-7474. [38] Kuttel M, Naidoo K J. Glycosidic linkage rotations determine amylose stretching mechanism [J]. Journal of the American Chemical Society, 2005, 127(1): 12-13. [39] Morris G M, Huey R, Lindstrom W, et al. AutoDock4 and AutoDockTools4: Automated docking with selective receptor flexibility [J]. Journal of computational chemistry, 2009, 30(16): 2785-2791. [40] Yee P, Shah J K, Maginn E J. State of hydrophobic and hydrophilic ionic liquids in aqueous solutions: are the ions fully dissociated?[J]. The Journal of Physical Chemistry B, 2013, 117(41): 12556-12566. [41] S.M. F, H. R, D. S, AutoDockTools, version 1.4. 5, The Scripps Research Institute, La Jolla, (2007) [42] Ravichandran S, Collins J R, Singh N, et al. A molecular model of the
enantioselective
liquid
chromatographic
separation
of
(R,
S)-ifosfamide and its N-dechloroethylated metabolites on a teicoplanin aglycon chiral stationary phase [J]. Journal of Chromatography A, 2012, 1269: 218-225. [43] Huang L, Lin J M, Yu L, et al. Improved simultaneous enantioseparation of β‐agonists in CE using β‐CD and ionic liquids [J]. Electrophoresis, 2009, 30(6): 1030-1036. [44] Jin Y, Chen C, Meng L, et al. Simultaneous and sensitive capillary electrophoretic
enantioseparation
of
three
β-blockers
with
the
combination of achiral ionic liquid and dual CD derivatives[J]. Talanta, 2012, 89: 149-154.
Table 1 Evaluation of the synergistic effect with CILs as additives NEF
μeof Analytes
(cm2s-1V-110-5)
Maltodextrin
KET
ECO
VOR
t1/ t2 (min)
Rs
α
t1/ t2 (min)
Rs
α
t1/ t2 (min)
Rs
α
t1/ t2 (min)
Rs
α
1.66
5.051/5.119
0.48
1.013
8.171/8.447
1.70
1.034
9.677/9.841
0.63
1.017
6.612/6.694
0.50
1.012
Maltodextrin+D-PAN-Na+
1.57
6.339/6.491
0.63
1.023
8.880/9.153
1.90
1.031
9.331/9.510
0.86
1.019
6.826/6.965
0.91
1.020
Maltodextrin+TMA-OH
-3.54
10.181/10.401
1.05
1.022
13.429/13.917
1.86
1.036
13.690/14.019
0.73
1.024
9.751/9.969
0.69
1.022
Maltodextrin+TMA-D-PAN
-4.10
14.655/15.155
1.87
1.041
17.572/19.453
4.91
1.107
18.089/18.821
2.18
1.041
16.254/16.776
2.49
1.032
Maltodextrin+TMA-D-QUI
-4.05
12.712/13.138
1.68
1.034
17.148/18.329
2.58
1.069
14.352/14.739
1.03
1.027
18.150/18.658
1.28
1.028
Conditions: 60 mM Tris-H3PO4 buffer solution containing 3.0% (w/v) maltodextrin and 50 mM D-PAN-Na+, 50 mM achiral IL (TMA-OH), 50 mM TMA-D-PAN or 50 mM TMA-D-QUI ; buffer pH 3.0; applied voltage, 24 kV (NEF), 26 kV (KET, ECO, VOR); capillary temperature, 20 °C, other conditions see section 2.
Table 2 Effects of TMA-D-PAN concentrations on the chiral separation of NEF, KET, ECO, VOR. TMA-D-PAN concentration ( mM ) 20 Analytes
30
40
50
60
t1/ t2 (min)
Rs
α
t1/ t2 (min)
Rs
α
t1/ t2 (min)
Rs
α
t1/ t2 (min)
Rs
α
t1/ t2 (min)
NEF
9.970/10.178
1.10
1.021
10.083/10.344
1.18
1.025
11.240/11.579
1.44
1.030
14.565/15.155
1.87
1.041
15.369/16.073
1.47
1.046
KET
12.085/12.871
2.98
1.065
13.564/14.493
3.56
1.069
15.367/16.480
3.78
1.072
17.572/19.453
4.91
1.107
21.959/23.358
3.97
1.064
ECO
13.015/13.301
1.13
1.023
13.369/13.730
1.21
1.027
15.219/15.828
1.61
1.030
18.089/18.821
2.18
1.041
21.042/21.780
1.86
1.035
VOR
11.143/11.410
0.97
1.024
12.623/12.965
1.02
1.027
14.868/16.776
1.35
1.032
16.254/16.776
2.49
1.032
20.210/20.911
Rs
1.76
Conditions: 60 mM Tris-H3PO4 buffer solution containing 3.0% (w/v) maltodextrin and 50 mM TMA-D-PAN; buffer pH 3.0; applied voltage, 24 kV (NEF), 26 kV (KET, ECO, VOR); capillary temperature, 20 °C, other conditions see section 2.
α
1.035
Table 3 Summary of molecular modeling with AutoDock NEF
KET
ECO
VOR
∆GRa
∆GSa
|∆∆G|a,b
∆GRa
∆GSa
|∆∆G|a,b
∆GRa
∆GSa
|∆∆G|a,b
∆GRa
∆GSa
|∆∆G|a,b
-5.46
-5.51
0.11
-7.42
-7.77
0.35
-5.94
-6.07
0.13
-5.29
-5.39
0.10
Maltodextrin/CIL1
-6.55
-6.78
0.23
-8.01
-8.48
0.47
-6.99
-7.26
0.27
-5.60
-5.90
0.30
Maltodextrin/CIL2
-6.16
-6.37
0.21
-7.69
-8.13
0.44
-6.77
-6.94
0.17
-5.45
-5.67
0.22
Maltodextrin/CIL1
-6.63
-7.04
0.41
-8.12
-8.88
0.76
-7.24
-7.68
0.39
-6.34
-6.79
0.35
Maltodextrin/CIL2
-6.49
-6.85
0.36
-7.93
-8.45
0.52
-6.97
-7.18
0.19
-5.80
-6.06
0.26
Single Maltodextrin Dissociated CILs
Associated CILs
CIL1: TMA-D-PAN; CIL2; TMA-D-QUI a Units are kcal/mol b Absolute value of different of free binding energies between the (R)-and (S)-enantiomer
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
S0021-9673(17)30840-3 http://dx.doi.org/doi:10.1016/j.chroma.2017.06.007 CHROMA 358577
To appear in:
Journal of Chromatography A
Received date: Revised date: Accepted date:
8-3-2017 26-5-2017 1-6-2017
Please cite this article as: Xuan Yang, Yingxiang Du, Zijie Feng, Zongran Liu, Jingtang Li, Establishment and molecular modeling study of Maltodextrinbased synergistic enantioseparation systems with two new hydroxy acid chiral ionic liquids as additives in capillary electrophoresis, Journal of Chromatography Ahttp://dx.doi.org/10.1016/j.chroma.2017.06.007 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Establishment and molecular modeling study of Maltodextrin-based synergistic enantioseparation systems with two new hydroxy acid chiral ionic liquids as additives in capillary electrophoresis Xuan Yang1, Yingxiang Du1, 2, 3, *, Zijie Feng1, Zongran Liu1, Jingtang Li1
1
Department of Analytical Chemistry, China Pharmaceutical University, Nanjing
210009, P. R. China 2
Key Laboratory of Drug Quality Control and Pharmacovigilance (Ministry of
Education), China Pharmaceutical University, Nanjing 210009, P. R. China 3
State Key Laboratory of Natural Medicines, China Pharmaceutical University,
Nanjing 210009, P. R. China
*
Correspondence: Professor Yingxiang Du, Department of Analytical Chemistry,
China Pharmaceutical University, No.24 Tongjiaxiang, Nanjing, Jiangsu 210009, China E-mail: [email protected] Tel./fax: +86 25 83221790
Highlights Two new chiral hydroxy acid-based ionic liquids (tertramethylammonium -D-pantothenate and tertramethylammonium-D-quinate) were designed and first used as additives to construct the maltodextrin-based synergistic system for enantioseparation in capillary electrophoresis (CE).
Compared to traditional system, significantly improved separations in the CIL/Maltodextrin synergistic systems were obtained.
The chiral recognition process of the synergistic systems based on maltodextrin was first evaluated by molecular modeling method.
The modeling results were in good agreement with our experimental results.
Abstract： Discovering more superior performance of ionic liquids for the separation science has triggered increasing interest. In this work, two new Hydroxy
acid-based
chiral
ionic
liquids
(tertramethylammonium-D-pantothenate
(CILs)
(TMA-D-PAN),
tertramethylammonium-D-quinate (TMA-D-QUI)) were designed and first used as additives to establish the maltodextrin-based synergistic systems for enantioseparation in capillary electrophoresis (CE). Compared to traditional single maltodextrin chiral separation system, significantly improved separations of
all tested drugs in the
CIL/Maltodextrin synergistic systems were obtained. Some parameters (CIL concentration, maltodextrin concentration, buffer pH, and applied voltage) in the TMA-D-PAN/Maltodextrin synergistic system have been examined and optimized for analytes. The molecular docking software AutoDock was applied to simulate the recognition process and surmise feasible resolution mechanism in the Maltodextrin/CILs synergistic systems, which has certain guiding value.
Keyword:
Chiral
ionic
liquid;
Synergistic
system;
electrophoresis; Enantioseparation; Molecular docking
Capillary
1. Introduction Chirality is one of the basic characteristics of nature. Almost half of the drugs in current use are known to be chiral and only a small number are administered as pure enantiomers. The popularity of the research on single enantiotopic chiral drugs stems from the fact that the desired pharmacological activity is exhibited by one enantiomer, while the other enantiomer may exhibit low activity, opposite pharmacokinetic and pharmacodynamics effects, or even toxicity. Among the various analytical techniques for enantioseparation, capillary electrophoresis (CE) has become a powerful alternative to chromatography, owing to its several advantages such as high resolving power, rapid analysis, low consumption of sample, solvent, and chiral selector, as well as high flexibility in choosing and changing types of selectors and separation modes[1]. The most common approaches for enantiomeric separation in CE involve the addition of one or more chiral selectors into the running buffer. Until now, various kinds of chiral selectors, including cyclodextrins and their derivatives[2-5], chiral crown ethers[6, 7], polysaccharides[8-10], antibiotics [11-13] and many other small molecules have been explored in CE for enantioseparation. However, in some cases, satisfactory results may not been obtained in the conventional separation systems using single chiral selector. Accordingly,
a number of research groups focus on hunting for different types of additives to enhance their enantioselectivity. Polysaccharides always possess variable structures and numerous functional chiral groups. Additionally, they have properties of the water-soluble and weak UV absorption, which benefit the application of these materials as CE chiral additives for enantioseparation.[9,10]. Maltodextrin, as an optically active polysaccharide chiral selector, can form the helical structure with hydrophilic outside and hydrophobic cavity in acidic and neutral aqueous solution[14].Compared with the rigid cavity of cyclodextrins, the spiral structure of maltodextrin, varying along with the size of the guest molecules, could be more flexible in chiral recognition[15,16].
However,
the
disadvantages
of
the
single
maltodextrin system (like high concentration, and finite enantioseparation capability) limit its application. Room-temperature ionic liquids (ILs), which are generally defined as organic salts in the liquid state under the condition of close to or below room temperature[17], have been developed fast in separation science, such as stationary phase in gas chromatography[18], mobile phase additives
in
liquid
chromatography[19],
solvent
in
two-phase
extraction[20]. In CE, ILs have been used as electrolyte[21, 22], running buffer modifiers[23], surface coating or chemical modification of the
capillary wall[24]. Chiral ionic liquids (CILs) are a subclass of ILs that have a either chiral cation, chiral anion, or both. On account of their potential chiral discrimination capabilities, the scientific interest is continuously increasing. However, in the recent years, only a few papers have reported the application of chiral ILs to establishment synergistic systems for enantioseparation in CE [25-30], and most of them are amino acid ionic liquids (AAILs) or their derivatives [26, 27]. As far as we know, only two papers about hydroxy acid ionic liquid as additives of the synergistic system [28, 29] were reported. In recent years, some studies have provided good insights into the host-guest interactions between CDs and the enantiomers and the chiral recognition mechanisms with molecular modeling methods[31-33], nuclear magnetic resonance (NMR)[34], X-ray crystallography[35]. Molecular modeling methods which included molecular dynamics simulations, Parametric Model 3(or 6) semiempirical methods, and molecular docking, have been performed to rationalize experimental findings. Moreover, molecular modeling methods have been successfully applied to evaluate the influence of hydrogen bonding in the chiral recognization process, which is difficult to explain because of the limitation of experimental methods. Investigations have never been reported which apply molecular modeling methods to elucidate the
possible chiral recognition mechanism of the polysaccharide-based synergistic systems. In
this
paper,
two
newly
tetramethylammonium-D-pantothenate tetramethylammonium-D-quinate
designed
hydroxy
acid
CILs
(TMA-D-PAN)
(TMA-D-QUI)
were
applied
and as
additives to evaluate their potential synergistic effect with maltodextrin as the chiral selector in CE. For the further understanding of the possible chiral recognition mechanism of synergistic system, we examined the course of host-guest inclusion between maltodextrin and enantiomers by means of a molecular docking technique and calculated the binding free energy by the AutoDock semi-empirical binding free energy function. The modeling results were in good agreement with our experimental results. 2. Experimental 2.1 Chemicals and reagents Maltodextrin (M040，DE4-7) was purchased from Sigma (St. Louis, Mo,
USA).Tetramethylammonium-D-pantothenate
Tetramethylammonium-D-quinate
(TMA-D-PAN),
(TMA-D-QUI)
and
Tetramethylammonium hydroxide (TMA-OH) ionic liquid (purity > 98%) were purchased from Shanghai Cheng Jie Chemical Co., Ltd. (Shanghai, China). Nefopan(NEF), Ketoconazole (KET), Econazole (ECO) and
Voriconazole(VOR), were supplied by Jiangsu Institute for Food and Drug Control (Nanjing, China). All these drug samples were racemic mixtures. The chemical structures of those substances are shown in Fig.1. Tris (hydroxymethyl) aminomethane (Tris) were purchased from Shanghai Huixing Biochemistry Reagent (Shanghai, China). Phosphoric acid and sodium hydroxide were of analytical regent from Nanjing Chemical Reagent (Nanjing, China). Double distilled water was used throughout all the experiments. Figure 1 2.2. Apparatus Electrophoretic experiments were performed with an Agilent 3D CE system (Agilent Technologies, Waldbronn, Germany), which consisted of a sampling device, a power supply, a photodiode array UV detector (wavelength range from 190 nm to 600 nm) and a date processor. The whole system was driven by Agilent ChemStation software (Revision B.02.01) for system control, date collection and analysis. It was equipped with a 50 cm (41.5 cm effective length) × 50 μm id uncoated fused-silica capillary (HebeiYongnian County Reafine Chromatography Ltd., Hebei, China). The samples were introduced hydrodynamically for 2 s (injection pressure 50 mbar). All separations were carried out at 20 oC using a voltage in the range of 20-28 kV. The wavelength for detection was 220
nm (NEF), 215 nm (KET, ECO, and VOR). The CE system was operated in the conventional mode with the anode at the injector end of the capillary. A new capillary was first rinsed with 1.0 M NaOH (30 min), followed by 0.1 M NaOH (20 min) and water (20 min), respectively. At the beginning of each day, the capillary was flushed with 0.1 M NaOH (10 min) followed by water (10 min). Between consecutive injections, the capillary was rinsed with 1.0 M NaOH, 0.1 M NaOH, water and running buffer for 3 min each. Nylon filters (0.45 μm) were purchased from Jiangsu Hanbon Science and Technology (Nanjing, China). All the solution need to be filtered before used. 2.3 Procedures The background electrolyte (BGE) consisted of 60 mM Tris solution (if not stated otherwise), adjusted to specified pH value with H 3PO4 (20%, v/v). The running buffer solutions were freshly prepared by dissolving appropriate amounts of maltodextrin and/or ILs in BGE, and then adjusting pH exactly to a desired value by adding a small volume of H3PO4 (20%, v/v) using a microsyringe. The racemic sample of NEF was dissolved
in
distilled
water.
The
racemic
samples
of
azole
antifungals(KET, ECO,VOR) were dissolved in methanol (50%，v/v).All those racemic samples were prepared at a concentration of 1.0 mg/ml.
Running buffers and samples were filtered with a 0.45 μm pore membrane filter and degassed by sonication prior to use. 2.4 Molecular Construction and Optimization Molecular construction was carried out using the ChemOffice 2010 software package. The MM2 force field and molecular dynamics were applied for the structure optimization. The PM3 quantum mechanical method was used for geometry optimization to adjust any spatially unreasonable bond distances and bond angles of all enantiomers. Glucose-α (1→4)-glucose is the backbone glycosidic linkage in maltodextrin [15,16]. The φ and ψ dihedral angles for the α(1→4)-linkage are define as φ = H1—C1—O1—C4＇, φ = C1—O1—C4＇—H4＇.The number of D-glucose units in the three-dimensional structure of the maltodextrin was set at 15 (based on the average of molecular weight)and the
crystal
structure
values
of
φ,ψ=5°,13°
in
the
vacuum
condition[36-38].Geometry optimization was conducted by applying the PM3 quantum mechanical method. 2.5 Molecular docking simulations Molecular docking simulations were performed with the automated docking program of AutoDock 4.2.3[39]. It employed the Lamacrkian genetic algorithm (LGA) to identify binding conformations of a flexible ligand (or small molecule) to a target receptor. The two CILs
(TMA-D-PAN and TMA-D-QUI)studied in this work were supposed to exist in two states of ionic associated and dissociated, resulting from the fact that ionic liquids are partially dissociated in the aqueous solutions due to the dual nature of ILs (hydrophilic/hydrophobic properties) associated to non-polar and ionic interactions[40].To comprehensive investigate the effect of the two chiral ionic liquids，associated CILs and dissociated CILs were separately introduced in the molecular model construction. As a preliminary preparation step to docking, AutoGrid[41] creates 3D grid boxes to generate a simplified representation for target receptor (e.g. maltodextrin). Each atom type (called probe) of the ligand will be placed at the grid points and its interaction energy with all the atoms of the receptor will be computed and assigned to the corresponding grid point[42]. Grid maps of dimensions70 A˚×60 A˚×60 A˚, with a grid spacing of 0.375 A˚, were placed to cover the maltodextrin molecule. One hundred LGA runs, each with 200 individuals in the population, were performed. Results differing by less than 1 A˚ in a positional root mean square deviation (rmsd) were clustered together. In each group, the lowest binding energy configuration with the highest % frequency was selected as the group representative. 3. Results and discussion 3.1 Effect of different chiral ILs and achiral IL
The electropherograms of four enantiomers (NEF, KET, ECO, and VOR) in different chiral separation systems was shown in Fig.2. Under the optimal conditions (60 mM Tris-H3PO4 buffer solution containing 3.0% (w/v) maltodextrin, 50 mM TMA-D-PAN or 50 mM TMA-D-QUI ; buffer pH 3.0; applied voltage, 24 kV for NEF, 26 kV for KET, ECO, VOR; capillary temperature, 20 °C), the entirely separations of all the tested drugs were obtained in the synergistic systems. As observed in Table 1, with the EOF reversed, the migration time of the analytes in ILs/Maltodextrin systems were longer than counterparts in the single maltodextrin system. This results from the increased ionic liquid strength of the running buffer, as well as the adsorption of IL cations (TMA+) on the capillary inner surface [43, 44]. The prolong migration time would provide more opportunities for chiral recognition in the separation process. A comparative study was conducted between achiral IL (TMA-OH)/Maltodextrin system and chiral ILs/Maltodextrin system. As observed, the addition of TMA-OH can also reverse the EOF in a similar extent. But the TMA-OH/Maltodextrin synergistic system exhibited poorer enantioseparation than the CILs/Maltodextrin systems, indicating the indispensability of the chiral parts of CILs in the synergistic systems. As excepted, significantly improved Rs and α of all the tested enantiomers were obtained in both of the TMA-D-PAN/Maltodextrin
system and TMA-D-QUI/Maltodextrin system, suggesting the existence of the synergistic effect between CILs and chiral selector maltodextrin. With regard to the two studied CILs, TMA-D-PAN exhibited better synergistic effect towards four analytes than TMA-D-QUI. It might be attributed to the chiral anion part of CILs, which is the only disparity between the structures of two CILs. This phenomenon indicated that the structure and properities of the chiral part of chiral ILs have a significant impact on the chiral discrimination process of the CILs synergistic systems [26-30]. It is worth noting that no enantioseparation for all tested drug were obtained with only CILs. To test the repeatability of the chiral separation systems, including single Maltodextrin, achiral IL/Maltodextrin, and CILs/Maltodextrin synergistic separation systems, five consecutive separations of four enantiomers were performed. For all studied analytes, the intra- and inter day variations of Rs (the RSD values of the Rs) were less than 3.6% and 3.9%, respectively. The intra- and interday variations of migration times (the RSD values of the migration times) were less than 3.3% and 3.7%， respectively. It could therefore be concluded that the synergistic systems could provide satisfactory repeatability. Table 1 Figure 2
3.2
Effect
of
CIL
concentration
on
enantioseparation
TMA-D-PAN/Maltodextrin synergistic system was selected to optimize the separation conditions. To investigate the effect of CIL concentration on enantioseparation, experiments were carried out by varying the CIL concentration in the range of 20 mM-60 mM while keeping maltodextrin concentration constant at 3.0% （w/v）. As observed in Table 2, the migration time of all the enantiomers increased with the CIL concentration changing from 20 mM to 60 mM mainly owing to a decrease in the density of negative charge resulting from the adsorption of the IL cations on the surface of the fused-silica capillary and, especially, the increased complexation (synergistic effect) between enantiomers and chiral recognition reagents. With the increase of migration time, the separation resolution and selectivity of NEF, KET, ECO, VOR, increased gradually with CIL concentration rising from 20 mM to 50 mM, indicating that the synergistic effect between chiral IL and maltodextrin continuously improved. At higher CIL concentration (50 mM-60 mM), the selectivity of KET, ECO decreased, that observation probably was owned to the saturation of the synergistic effect between CIL and maltodextrin in the concentration of 50 mM. While the selectivity of NEF, VOR continued to increase in 60 mM, the separation
resolution declined presumably, due to the minute broadening peak at high CILs concentration. Taking account of good separation and appropriate migration time simultaneously, the CIL concentration of 50 mM was determined to optimum for the synergistic system. Table 2 3.3 Effect of maltodextrin concentration on enantioseparation The concentration of chiral selectors is also an important factor influencing the enantioseparation. To obtain the optimal selector concentration of the TMA-D-pantothenate/Maltodextrin synergistic system, we investigated a series of concentrations (1.5%-3.5% (w/v)) of Maltodextrin with 60 mM Tris/H3PO4 buffer solution (pH 3.0) containing 50 mM TMA-D-pantothenate. At the same time, we studied the single maltodextrin system, and all studied drugs were partially separated. As shown in Fig.3, the Rs and α of all the racemic drugs initially increased along with concentration of chiral selector increasing from 1.5% (w/v) to 3.0% (w/v), which was probably imputed to the change of solution viscosity and the enhancement of interaction among maltodextrin, CIL and enantiomers. Then the Rs and α of three chiral pharmaceutical (NEF, KET, ECO) decreased at high concentration (3.5% (w/v)), indicating the oversaturated situation of the synergistic system. The selectivity of VOR
continuously increased, while the resolution declined mainly due to the fact that it appeared to sustain the peaking broadening. Moreover, compared with the single maltodextrin system, TMA-A-PAN/Maltodextrin synergistic system could achieve better separation resolution under all the experimental conditions of maltodextrin concentration. By the results, 3.0% (w/v) maltodextrin was selected for further analysis. Figure 3 3.4 Effect of buffer pH on enantioseparation Buffer
pH
is
a
critical
and
sensitive
parameter
for
the
CIL/Maltodextrin synergistic system optimization due to the significant influence in the ionic state of capillary wall, the dissociation of analytes, the electrophoretic behavior of solutes and the interaction among chiral selectors, chiral ILs and enantiomers (e.g. electrostatic interaction and hydrogen bondings). The effect of the BGE pH on chiral separation was investigated by increasing the pH from 2.0 to 4.0 using 60 mM Tris/H3PO4 containing 3.0% (w/v) maltodextrin and 50 mM TMA-Dpantothenate, while all the analytes with a positive charge. As can be seen from Fig.4, the separation resolutions and selectivities for all the chiral drugs increased when pH increased from 2.0 to 3.0 indicating the enhancement of the synergistic system effect, and then they decreased
when pH further increased from 3.0 to 4.0. This observation could be explained by the fact that the enhancement of the synergistic system effect with the pH increased, thus chiral resolution and selectivity was increased. However, further increasing of pH would mainly weaken the interaction among chiral selectors, chiral ILs and enantiomers. Therefore, buffer pH at 3.0 was selected for further analysis. Figure 4 3.5 Effect of applied voltage on enantioseparation In CE, applied voltage is an important factor to control the column efficiency, the resolution, and the analysis time. Higher voltage would bring about higher CE efficiency and shorter migration time, while it leaded to encumbered Joule heating, which mainly reduced the resolution. In another way, the prolonged migration time along with lower applied voltage would give more chances for the enantiorecognition, which always influences the resolution. In order to evaluate the effect of applied voltage on chiral separation of the synergistic system, we studied a series of voltage (20 kV-28 kV) with 60 mM Tris/H3PO4 buffer solution (pH 3.0) containing 50mM TMA-D-pantothenate and 3% (w/v) maltodextrin. As shown in Fig.5, the resolutions of all racemic drugs initially increased owning to the improvement of CE efficiency with the applied voltage increasing, and
then the decline of resolution appeared probably because of extra Joule heating and peaking broadening. Considering enantioseparation and migration time, we selected applied voltage of 24 kV for NEF and 26 kV for KET, ECO, VOR in the synergistic system. Figure 5 4. Molecular modeling study of chiral recognition mechanism In this study, we used the molecular docking software AutoDock 4.2.3 to confirm and explain the possible chiral recognition mechanism and the potential synergistic effect of the CILs/Maltodextrin synergistic system. The molecular docking configurations for KET enantiomers in single maltodextrin synergistic
system system
(b)
(a), and
dissociated-TMA-D-PAN/Maltodextrin associated-TMA-D-PAN/Maltodextrin
synergistic system (c) are presented in Fig.6. Generally, there are multiple interactions participating in the process of the enantiorecognition, including hydrogen bonding interaction, dipole-dipole, π-π, charge-charge, host-guest inclusion, etc. Although not all the interactions could be observed directly, the molecular docking configurations of the inclusion complexes still provide visualization of some interactions for the investigation of chiral recognition mechanism. As shown in Fig.6, hydrogen bonding interactions (labeled by the green line) for KET were dramatically enhanced in TMA-D-PAN/Maltodextrin synergistic systems
(no matter what state of TMA-D-PAN is) compared with single maltodextrin system. Meanwhile, the σ-π binding interaction (indicated by orange line) appeared between enantiomer and maltodextrin in associated TMA-D-PAN/Maltodextrin synergistic system. The results above suggested the CILs truly have impact on the recognition of maltodextrin. The molecular docking results with AutoDock for all tested enantiomers in both single maltodextrin and CILs/Maltodextrin synergistic systems are presented in Table 3. Thermodynamically, the more negative of binding energies indicates the more stable state of the inclusions. As shown in Table 3, whatever the state of CILs is, the addition of CILs to the single maltodextrin system improved the values of |∆G| for all studied drugs. Moreover, the results showed that the participation of dissociated and associated CILs could increase the difference of binding free energies (|∆∆G|) between enantiomers significantly, which might be attributed to the increased chiral discrimination and improved chiral selectivity in maltodextrin separation systems. Simultaneously, the difference of the calculated binding energy corresponded with the experimental chiral selectivity α (see Table 1). In addition, the values of |∆∆G| in the associated CILs/Maltodextrin system were higher than those in the dissociated CILs/Maltodextrin system for
all four enantiomers, indicating the essential role of associated CILs in CILs/Maltodextrin synergistic system for chiral separation. Above all, the existence of the CILs may ameliorate the chiral environment, and thus improve the chiral recognition ability and selectivity of the original system, indicating the synergistic effect in the CILs/Maltodextrin. Figure 6 Table 3
5. Conclusions In this paper, two novel chiral ILs based on hydroxyl acid ， TMA-D-PAN and TMA-D-QUI, were first used as additives to establish the Maltodextrin-based synergistic systems in CE for enantioseparation. Some parameters (CIL concentration, maltodextrin concentration, buffer pH, and applied voltage) in the TMA-D-PAN/Maltodextrin synergistic system have been examined and optimized for all tested drugs. Significant improvements of chiral separation for analytes were obtained with CILs compared to the single maltodextrin system. The chiral recognition mechanism of the CILs/Maltodextrin synergistic systems were further investigated with the molecular docking software AutoDock 4.2.3., which serves as a good method for predicting enantioseparation.
Acknowledgements This work was supported by the Project of National Natural Science Foundation of China (No.: 81373378) and the Natural Science Foundation of Jiangsu Province (Program No.BK20150697) and the Project of the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD).
The authors have declared no conflict of interst
Figure Caption
Fig.1 Structures of (A) Maltodextrin (B) TMA-OH (C) TMA-D-PAN (CIL1) and TMA-D-QUI (CIL2) (D) Four racemic drugs.
Fig.2 Typical electrophoregrams of the chiral separation of (A) NEF, (B) KET, (C) ECO, (D) VOR in the different separation system. Conditions: 60 mM Tris-H3PO4 buffer solution containing (a) 3.0% (w/v) maltodextrin; (b) 3.0% (w/v) maltodextrin and 50 mM D-PAN-Na+; (c) 3.0% (w/v) maltodextrin and 50 mM TMA-OH; (d) 3.0% (w/v) maltodextrin and 50 mM TMA-D-PAN; (e) 3.0% (w/v) maltodextrin and 50 mM TMA-D-QUI; buffer pH 3.0; applied voltage, 24 kV (NEF), 26 kV (KET, ECO, VOR); capillary temperature, 20 oC other condition see section 2.
Fig.3 Effect of chiral selector concentration on the enantioseparation. Conditions: 60 mM Tris-H3PO4 buffer solution (pH 3.0) containing 1.5%-3.5% (w/v) maltodextrin with 50 mM CIL1 (TMA-D-PAN); applied voltage, 24 kV (NEF), 26 kV (KET, ECO, VOR); capillary temperature, 20 °C; other condition see section 2.
Fig.4 Effect of buffer pH on the enantioseparation. Conditions: 60 mM
Tris-H3PO4 buffer solution containing 3.0% (w/v) maltodextrin and 50 mM TMA-D-PAN; buffer pH 2.0-4.0; applied voltage, 24 kV (NEF), 26 kV (KET, ECO, VOR); capillary temperature, 20 °C; other condition see section 2.
Fig.5 Effect of applied voltage on the enantioseparation. Conditions: 60 mM Tris-H3PO4 buffer solution containing 3.0% (w/v) maltodextrin and 50 mM TMA-D-PAN; buffer pH 3.0; applied voltage, 20 kV-28 kV; capillary temperature, 20 °C; other condition see section 2.
Fig. 6 Molecular docking configuration for KET enantioseparation in a single Maltodextrin system; b dissocated TAM-D-PAN/Maltoextrin synergistic
system
and
c
associated
TMA-D-PAN/Maltodextrin
synergistic system. The left is (R)-enantiomer and the right is (S)-enantiomer. The hydrogen bonding is indicated by green dotted line. The σ-π is indicated by orange line. O red，N blue，H white，C grey.
References [1] Koppenhoefer B, Epperlein U, Christian B, et al. Separation of enatiomers of drugs by capillary electrophoresis III. β-cyclodextrin as chiral solvating agent [J]. Journal of Chromatography A, 1996, 735(1-2): 333-343. [2]Saz J M, Marina M L. Recent advances on the use of cyclodextrins in the chiral analysis of drugs by capillary electrophoresis [J]. Journal of Chromatography A, 2016, 1467: 79-94. [3] Zhu Q, Scriba G K E. Advances in the use of cyclodextrins as chiral selectors in capillary electrokinetic chromatography: fundamentals and applications [J]. Chromatographia, 2016, 79(21-22): 1403-1435. [4] Řezanka P, Navrátilová K, Řezanka M, et al. Application of cyclodextrins in chiral capillary electrophoresis [J]. Electrophoresis, 2014, 35(19): 2701-2721. [5] Escuder-Gilabert L, Martín-Biosca Y, Medina-Hernández M J, et al. Cyclodextrins in capillary electrophoresis: recent developments and new trends [J]. Journal of Chromatography A, 2014, 1357: 2-23. [6]Paik M J, Kang J S, Huang B S, et al. Development and application of chiral crown ethers as selectors for chiral separation in high-performance liquid chromatography and nuclear magnetic resonance spectroscopy [J]. Journal of Chromatography A, 2013, 1274: 1-5. [7] Kuhn R. Enantiomeric separation by capillary electrophoresis using a
crown ether as chiral selector [J]. Electrophoresis, 1999, 20(13): 2605-2613. [8] Nishi H, Izumoto S, Nakamura K, et al. Dextran and dextrin as chiral selectors in capillary zone electrophoresis [J]. Chromatographia, 1996, 42(11): 617-630. [9] Chen J, Du Y, Zhu F, et al. Glycogen: a novel branched polysaccharide chiral selector in CE [J]. Electrophoresis, 2010, 31(6): 1044-1050. [10] Zhang Q, Du Y, Chen J, et al. Investigation of chondroitin sulfate D and chondroitin sulfate E as novel chiral selectors in capillary electrophoresis [J]. Analytical and bioanalytical chemistry, 2014, 406(5): 1557-1566. [11]Chen B, Du Y, Wang H. Study on enantiomeric separation of basic drugs by NACE in methanol ‐ based medium using erythromycin lactobionate as a chiral selector[J]. Electrophoresis, 2010, 31(2): 371-377. [12] Domínguez‐Vega E, Pérez‐Fernández V, Crego A L, et al. Recent advances in CE analysis of antibiotics and its use as chiral selectors [J]. Electrophoresis, 2014, 35(1): 28-49. [13]Domínguez‐Vega E, Montealegre C, Marina M L. Analysis of antibiotics by CE and their use as chiral selectors: An update [J]. Electrophoresis, 2016, 37(1): 189-211. [14] Nishi H, Izumoto S, Nakamura K, et al. Dextran and dextrin as chiral
selectors in capillary zone electrophoresis[J]. Chromatographia, 1996, 42(11): 617-630. [15] Stefan R I, Van Staden J F, Aboul-Enein H Y, et al. Maltodextrins as new chiral selectors in the design of potentiometric, enantioselective membrane electrodes[J]. Fresenius' journal of analytical chemistry, 2001, 370(1): 33-37. [16] Tabani H, Fakhari A R, Nojavan S. Maltodextrins as chiral selectors in CE: Molecular structure effect of basic chiral compounds on the enantioseparation[J]. Chirality, 2014, 26(10): 620-628. [17] Tang S, Liu S, Guo Y, et al. Recent advances of ionic liquids and polymeric ionic liquids in capillary electrophoresis and capillary electrochromatography[J]. Journal of Chromatography A, 2014, 1357: 147-157. [18] Zhang C, Park R A, Anderson J L. Crosslinked structurally-tuned polymeric ionic liquids as stationary phases for the analysis of hydrocarbons
in
kerosene
and
diesel
fuels
by comprehensive
two-dimensional gas chromatography [J]. Journal of Chromatography A, 2016, 1440: 160-171. [19] Wang Q, Chen X, Qiu B, et al. Ionic liquid as a mobile phase additive in high ‐ performance liquid chromatography for the simultaneous determination of eleven fluorescent whitening agents in paper materials[J]. Journal of Separation Science, 2016, 39(7):1242.
[20]Tang F, Zhang Q, Ren D, et al. Functional amino acid ionic liquids as solvent and selector in chiral extraction [J]. Journal of Chromatography A, 2010, 1217(28): 4669-4674. [21] Zhang H, Qi L, Mu X, et al. Influence of ionic liquids as electrolyte additives on chiral separation of dansylated amino acids by using Zn (II) complex mediated chiral ligand exchange CE[J]. Journal of separation science, 2013, 36(5): 886-891. [22] Liu R, Du Y, Chen J, et al. Investigation of the enantioselectivity of tetramethylammonium L-hydroxyproline ionic liquid as a novel chiral ligand in ligand-exchange CE and ligand-exchange MEKC[J]. Chirality, 2015, 27(1): 58-63. [23]Borissova M, Palk K, Koel M. Micellar electrophoresis using ionic liquids [J]. Journal of Chromatography A, 2008, 1183(1): 192-195. [24]Jin Y, Chen C, Meng L, et al. Simultaneous and sensitive capillary electrophoretic
enantioseparation
of
three
β-blockers
with
the
combination of achiral ionic liquid and dual CD derivatives[J]. Talanta, 2012, 89: 149-154. [25] François Y, Varenne A, Juillerat E, et al. Evaluation of chiral ionic liquids as additives to cyclodextrins for enantiomeric separations by capillary electrophoresis[J]. Journal of Chromatography A, 2007, 1155(2): 134-141. [26] Zhang J, Du Y, Zhang Q, et al. Investigation of the synergistic effect
with amino acid-derived chiral ionic liquids as additives for enantiomeric separation in capillary electrophoresis [J]. Journal of Chromatography A, 2013, 1316: 119-126. [27] Zhang Q, Du Y. Evaluation of the enantioselectivity of glycogen-based synergistic system with amino acid chiral ionic liquids as additives in capillary electrophoresis [J]. Journal of Chromatography A, 2013, 1306: 97-103. [28] Zuo L, Meng H, Wu J, et al. Combined use of ionic liquid and β‐ CD for enantioseparation of 12 pharmaceuticals using CE [J]. Journal of separation science, 2013, 36(3): 517-523 [29] Cui Y, Ma X, Zhao M, et al. Combined Use of Ionic Liquid and Hydroxypropyl ‐ β ‐ Cyclodextrin for the Enantioseparation of Ten Drugs by Capillary Electrophoresis [J]. Chirality, 2013, 25(7): 409-414. [30] Zhang Y, Du S, Feng Z, et al. Evaluation of synergistic enantioseparation systems with chiral spirocyclic ionic liquids as additives by capillary electrophoresis[J]. Analytical and bioanalytical chemistry, 2016, 408(10): 2543-2555. [31] Li W, Liu C, Tan G, Zhang X, Zhu Z, Chai Y, Molecular modeling study of chiral separation and recognition mechanism of beta-adrenergic antagonists by capillary electrophoresis, Int. J. Mol. Sci, 13 (2012) 710-725. [32] Alvira E, Molecular dynamics study of the influence of solvents on
the chiral discrimination of alanine enantiomers by β-cyclodextrin, Tetrahedron: Asymmetry, 24 (2013) 1198-1206. [33] Nascimento C S, Lopes J F, Guimarães L, et al. Molecular modeling study of the recognition 4-hydroxypropranolol
mechanism and
by
capillary
enantioseparation of
electrophoresis
using
carboxymethyl-β-cyclodextrin as the chiral selector[J]. Analyst, 2014, 139(16): 3901-3910. [34] Li W, Tan G, Zhao L, et al. Computer-aided molecular modeling study of enantioseparation of iodiconazole and structurally related triadimenol analogues by capillary electrophoresis: chiral recognition mechanism and mathematical model for predicting chiral separation[J]. Analytica chimica acta, 2012, 718: 138-147. [35] Stiufiuc R, Iacovita C, Stiufiuc G, et al. Surface mediated chiral interactions between cyclodextrins and propranolol enantiomers: a SERS and DFT study[J]. Physical Chemistry Chemical Physics, 2015, 17(2): 1281-1289. [36] Quigley G J, Sarko A, Marchessault R H. Crystal and molecular structure of maltose monohydrate [J]. Journal of the American Chemical Society, 1970, 92(20): 5834-5839. [37] Kuttel M M, Naidoo K J. Free energy surfaces for the α (1→ 4)-glycosidic linkage: implications for polysaccharide solution structure and dynamics [J]. The Journal of Physical Chemistry B, 2005, 109(15):
7468-7474. [38] Kuttel M, Naidoo K J. Glycosidic linkage rotations determine amylose stretching mechanism [J]. Journal of the American Chemical Society, 2005, 127(1): 12-13. [39] Morris G M, Huey R, Lindstrom W, et al. AutoDock4 and AutoDockTools4: Automated docking with selective receptor flexibility [J]. Journal of computational chemistry, 2009, 30(16): 2785-2791. [40] Yee P, Shah J K, Maginn E J. State of hydrophobic and hydrophilic ionic liquids in aqueous solutions: are the ions fully dissociated?[J]. The Journal of Physical Chemistry B, 2013, 117(41): 12556-12566. [41] S.M. F, H. R, D. S, AutoDockTools, version 1.4. 5, The Scripps Research Institute, La Jolla, (2007) [42] Ravichandran S, Collins J R, Singh N, et al. A molecular model of the
enantioselective
liquid
chromatographic
separation
of
(R,
S)-ifosfamide and its N-dechloroethylated metabolites on a teicoplanin aglycon chiral stationary phase [J]. Journal of Chromatography A, 2012, 1269: 218-225. [43] Huang L, Lin J M, Yu L, et al. Improved simultaneous enantioseparation of β‐agonists in CE using β‐CD and ionic liquids [J]. Electrophoresis, 2009, 30(6): 1030-1036. [44] Jin Y, Chen C, Meng L, et al. Simultaneous and sensitive capillary electrophoretic
enantioseparation
of
three
β-blockers
with
the
combination of achiral ionic liquid and dual CD derivatives[J]. Talanta, 2012, 89: 149-154.
Table 1 Evaluation of the synergistic effect with CILs as additives NEF
μeof Analytes
(cm2s-1V-110-5)
Maltodextrin
KET
ECO
VOR
t1/ t2 (min)
Rs
α
t1/ t2 (min)
Rs
α
t1/ t2 (min)
Rs
α
t1/ t2 (min)
Rs
α
1.66
5.051/5.119
0.48
1.013
8.171/8.447
1.70
1.034
9.677/9.841
0.63
1.017
6.612/6.694
0.50
1.012
Maltodextrin+D-PAN-Na+
1.57
6.339/6.491
0.63
1.023
8.880/9.153
1.90
1.031
9.331/9.510
0.86
1.019
6.826/6.965
0.91
1.020
Maltodextrin+TMA-OH
-3.54
10.181/10.401
1.05
1.022
13.429/13.917
1.86
1.036
13.690/14.019
0.73
1.024
9.751/9.969
0.69
1.022
Maltodextrin+TMA-D-PAN
-4.10
14.655/15.155
1.87
1.041
17.572/19.453
4.91
1.107
18.089/18.821
2.18
1.041
16.254/16.776
2.49
1.032
Maltodextrin+TMA-D-QUI
-4.05
12.712/13.138
1.68
1.034
17.148/18.329
2.58
1.069
14.352/14.739
1.03
1.027
18.150/18.658
1.28
1.028
Conditions: 60 mM Tris-H3PO4 buffer solution containing 3.0% (w/v) maltodextrin and 50 mM D-PAN-Na+, 50 mM achiral IL (TMA-OH), 50 mM TMA-D-PAN or 50 mM TMA-D-QUI ; buffer pH 3.0; applied voltage, 24 kV (NEF), 26 kV (KET, ECO, VOR); capillary temperature, 20 °C, other conditions see section 2.
Table 2 Effects of TMA-D-PAN concentrations on the chiral separation of NEF, KET, ECO, VOR. TMA-D-PAN concentration ( mM ) 20 Analytes
30
40
50
60
t1/ t2 (min)
Rs
α
t1/ t2 (min)
Rs
α
t1/ t2 (min)
Rs
α
t1/ t2 (min)
Rs
α
t1/ t2 (min)
NEF
9.970/10.178
1.10
1.021
10.083/10.344
1.18
1.025
11.240/11.579
1.44
1.030
14.565/15.155
1.87
1.041
15.369/16.073
1.47
1.046
KET
12.085/12.871
2.98
1.065
13.564/14.493
3.56
1.069
15.367/16.480
3.78
1.072
17.572/19.453
4.91
1.107
21.959/23.358
3.97
1.064
ECO
13.015/13.301
1.13
1.023
13.369/13.730
1.21
1.027
15.219/15.828
1.61
1.030
18.089/18.821
2.18
1.041
21.042/21.780
1.86
1.035
VOR
11.143/11.410
0.97
1.024
12.623/12.965
1.02
1.027
14.868/16.776
1.35
1.032
16.254/16.776
2.49
1.032
20.210/20.911
Rs
1.76
Conditions: 60 mM Tris-H3PO4 buffer solution containing 3.0% (w/v) maltodextrin and 50 mM TMA-D-PAN; buffer pH 3.0; applied voltage, 24 kV (NEF), 26 kV (KET, ECO, VOR); capillary temperature, 20 °C, other conditions see section 2.
α
1.035
Table 3 Summary of molecular modeling with AutoDock NEF
KET
ECO
VOR
∆GRa
∆GSa
|∆∆G|a,b
∆GRa
∆GSa
|∆∆G|a,b
∆GRa
∆GSa
|∆∆G|a,b
∆GRa
∆GSa
|∆∆G|a,b
-5.46
-5.51
0.11
-7.42
-7.77
0.35
-5.94
-6.07
0.13
-5.29
-5.39
0.10
Maltodextrin/CIL1
-6.55
-6.78
0.23
-8.01
-8.48
0.47
-6.99
-7.26
0.27
-5.60
-5.90
0.30
Maltodextrin/CIL2
-6.16
-6.37
0.21
-7.69
-8.13
0.44
-6.77
-6.94
0.17
-5.45
-5.67
0.22
Maltodextrin/CIL1
-6.63
-7.04
0.41
-8.12
-8.88
0.76
-7.24
-7.68
0.39
-6.34
-6.79
0.35
Maltodextrin/CIL2
-6.49
-6.85
0.36
-7.93
-8.45
0.52
-6.97
-7.18
0.19
-5.80
-6.06
0.26
Single Maltodextrin Dissociated CILs
Associated CILs
CIL1: TMA-D-PAN; CIL2; TMA-D-QUI a Units are kcal/mol b Absolute value of different of free binding energies between the (R)-and (S)-enantiomer
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6