Chiral Separation Techniques Research Articles

Simultaneous formation of helical and sheet-like assemblies from short azapeptides enables spontaneous resolution.

As determined by the homochirality of amino acid building units, protein secondary structures α-helix and β-sheet are single-handed chiral superstructures extending in one and quasi-two dimensions, respectively. Synthetic molecular assemblies that mimic the structural homochirality of proteins would provide insights into the origin of biological homochirality and inform the development of chiral separation techniques. Here we fabricated a homochiral 3D assembly consisting of 1D helical and 2D sheet-like assemblies that feature molecular packings resembling α-helix and β-sheet, respectively. This was achieved by using an alanine derivative, a β-turn structured short azapeptide from p-iodobenzoylalanine-based N-amido-N'-phenylthiourea. While N-H⋯SC/OC hydrogen bonds between the β-turn scaffolds afford a 2D pleated sheet-like structure, the head-to-tail C-I⋯π halogen bonds, together with the N-H⋯OC hydrogen bonds, support a 1D helical-like assembly, serving as linkers to connect the 2D sheet-like structures into a 3D superstructure. The two biomimetic assembly modes share the N-H⋯OC hydrogen bonds and can allow 3D homochiral elongation, driving spontaneous resolution of the short azapeptides to generate conglomerate crystals.

Kinetic resolution of substituted amido[2.2]paracyclophanes via asymmetric electrophilic amination

Planar chiral [2.2]paracyclophane derivatives are a type of structurally intriguing and practically useful chiral molecules, which have found a range of important applications in the field of asymmetric catalysis and material science. However, access to enantioenriched [2.2]paracyclophanes represents a longstanding challenge in organic synthesis due to their unique structures, which are still highly dependent on the chiral chromatography separation technique and classical chemical resolution strategy to date. In this work, we report on an efficient and versatile kinetic resolution protocol for various substituted amido[2.2]paracyclophanes, including those with pseudo-geminal, pseudo-ortho, pseudo-meta and pseudo-para disubstitutions, using chiral phosphoric acid (CPA)-catalyzed asymmetric amination reaction, which was also applicable to the enantioselective desymmetrization of an achiral diamido[2.2]paracyclophane. Detailed experimental studies shed light on a new reaction mechanism for the electrophilic aromatic C-H amination, which proceeded through sequential triazane formation and N[1,5]-rearrangement. The facile large-scale kinetic resolution reaction and diverse derivatizations of both the recovered chiral starting materials and the C-H amination products showcased the potential of this method.

Chiral separation and molecular modeling study of decursinol and its derivatives using polysaccharide-based chiral stationary phases

Plant-based bioactive substances have long been used to treat inflammatory ailments, owing to their low toxicity and cost-effectiveness. To enhance plant treatment by eliminating undesirable isomers, optimizing the chiral separation techniques in pharmaceutical and clinical studies is important. This study reported a simple and effective method for chiral separation of decursinol and its derivatives, which are pyranocoumarin compounds with anti-cancer and anti-inflammatory properties. Baseline separation (Rs >1.5) was achieved using five different polysaccharide-based chiral stationary phases (CSPs) that differed in chiral origin, chiral selector chemistry, and preparation technique. To separate all six enantiomers simultaneously, n-hexane and three alcohol modifiers (ethanol, isopropanol, and n-butanol) were used as mobile phases in the normal-phase mode. The chiral separation ability of each column with various mobile phase compositions was compared and discussed. As a result, amylose-based CSPs with linear alcohol modifiers demonstrated superior resolution. Three cases of elution order reversal caused by modifications of CSPs and alcohol modifiers were observed and thoroughly analyzed. To elucidate the chiral recognition mechanism and enantiomeric elution order (EEO) reversal phenomenon, detailed molecular docking simulations were conducted. The R- and S-enantiomers of decursinol, epoxide, and CGK012 exhibited binding energies of –6.6, –6.3, –6.2, –6.3, –7.3, and –7.5 kcal/mol, respectively. The magnitude of the difference in binding energies was consistent with the elution order and enantioselectivity (α) of the analytes. The molecular simulation results demonstrated that hydrogen bonds, π–π interactions, and hydrophobic interactions have a significant impact on chiral recognition mechanisms. Overall, this study presented a novel and logical approach of optimizing chiral separation techniques in the pharmaceutical and clinical industries. Our findings could be further applied for screening and optimizing enantiomeric separation.

Preparation of a cyclodextrin metal-organic framework (CD-MOF) membrane for chiral separation

Due to their unique properties, the efficient separation of racemic mixtures remains a great challenge. Recently, membrane separation has been demonstrated as a promising chiral separation technique with the advantages of continuous operation, high throughput, and cost effectiveness. In this work, a cyclodextrin-metal-organic framework (CD-MOF) was prepared and then composited with polyvinylidene fluoride (PVDF) polymer by the phase inversion via immersion precipitation method to form a CD-MOF/PVDF composite membrane for chiral amino acid separation for the first time. The composite membranes prepared under different conditions were characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR) thermogravimetric analysis (TGA)and differential scanning calorimetry (DSC). The effects of preparation conditions including CD-MOF load, polymer concentration, and casting solution temperature on the hydrophilicity, water flux, bovine serum albumin (BSA) rejection and chiral separation performance of the membrane were investigated. It is found that the composite membrane (V-5%−12) prepared with the CD-MOF load of 5%, polymer concentration of 12%, and casting solution of 70 ℃ shows the best performance with the water flux of 466.4 L/m2∙h, BSA rejection rate of 24.3% and stable 100% separation efficiency for phenylalanine (Phe) enantiomers for 4 h. The interaction energies between the CD-MOF and D-Phe and L-Phe were calculated using the Gaussian 09 program package and the chiral separation mechanism of the composite membrane was proposed at the molecular level. Our work demonstrates great development potentials of the CD-MOF/PVDF composite membrane in the chiral separation of drugs.

Dynamic Diastereomerism on Chiral Surfaces.

Adsorption of chiral molecules on chiral surfaces implies diastereomerism, evident in the adoption of distinct adsorption geometries. We show here that this diastereomerism produces a signature in the motion of chiral molecules desorbing from a chiral surface. The rotations of S- and R-alanine molecules are analyzed upon desorption from R-Cu{531} using first-principles molecular dynamics simulations. S-Ala molecules exhibit a larger angular momentum, with a clear preference for one rotational sense, whereas no such preference is observed for R-Ala molecules upon desorption from this surface. These trends would be reversed for desorption from the S-Cu{531} surface. Possible applications include chiral separation techniques and enantiospecific sensors.

Spherical core-shell Sb@C for tartaric acid enantioseparation

Enantiomers often exhibit different even opposite physiological or pharmacological activities, so it is necessary to develop chiral separation techniques to obtain single chiral enantiomers for further study and utilization of their physical and chemical properties. Herein, a simple one-pot method to synthesize Sb@C enantioseparation material with a controllable spherical core–shell structure was proposed, and the formation mechanism of the core–shell structure was clarified. Furthermore, capping of tartaric acid enantiomer on the Sb-S containing substance depending on complexation was verified an effective solution to design materials with enantioselectivity and the possible separation mechanism by hydrogen bonding based on sterical matching was also revealed. In addition, structure and enantioseparation capacity of the Sb@C materials are controllable by adjusting the ratio of Sb/S. The highest enantiomer separation capacity was up to ee = 25.80 % in all the Sb@C materials. This work presented a feasible and relatively universal strategy to design enantioseparation materials with potential application.

Chiral stationary phases and applications in gas chromatography.

Chiral compounds are ubiquitous in nature and play a pivotal role in biochemical processes, in chiroptical materials and applications, and as chiral drugs. The analysis and determination of the enantiomeric ratio (er) of chiral compounds is of enormous scientific, industrial, and economic importance. Chiral separation techniques and methods have become indispensable tools to separate chiral compounds into their enantiomers on an analytical as well on a preparative level to obtain enantiopure compounds. Chiral gas chromatography and high-performance liquid chromatography have paved the way and fostered several research areas, that is, asymmetric synthesis and catalysis in organic, medicinal, pharmaceutical, and supramolecular chemistry. The development of highly enantioselective chiral stationary phases was essential. In particular, the elucidation and understanding of the underlying enantioselective supramolecular separation mechanisms led to the design of new chiral stationary phases. This review article focuses on the development of chiral stationary phases for gas chromatography. The fundamental mechanisms of the recognition and separation of enantiomers and the selectors and chiral stationary phases used in chiral gas chromatography are presented. An overview over syntheses and applications of these chiral stationary phases is presented as a practical guidance for enantioselective separation of chiral compound classes and substances by gas chromatography.

Nafion membranes modified by cationic cyclodextrin derivatives for enantioselective separation

Nafion117® membranes modified by three cationic cyclodextrin (CD) derivatives have been prepared by strong ionic bonding. All CD derivatives contained bis(methylimidazolium) (MIM2) cationic anchor covalently bound to the CD unit, either using no spacer or using diethylene glycol (DEG) or tetraethylene glycol (TEEG) spacers. The modified membranes were tested in chiral separation of a model racemic mixture (d/l-tryptophan) from water. Different experimental set-ups for characterising membranes in enantioselective separation – pertraction, two kinds of sorption, and pressure-driven membrane separation – have been described and rigorously compared. The membranes CD-MIM2, CD-DEG-MIM2 have reached the highest enantiomeric excess, 14 and 44% respectively, in 280 days. The lowest performance of the CD-TEEG-MIM2 membrane, with the long spacer, has been visibly ameliorated by applying pertraction; enantiomeric excess rose from 2 to 27% in 80 days. Even though sorption played the main role in pertraction, this process substantially enhanced the separation of racemic mixtures. The pressure-driven approach has allowed the operation to be continuous and faster, which has the potential for continuous large-scale production of enantiopure compounds and could pave the way for many more commercial applications, satisfying the considerable demand for large-scale chiral separation techniques.

Novel optically active 2D materials based on λ-zirconium phosphate and chiral monocarboxylic acids: Synthesis and characterization

Five novel chiral 2D materials with general formula λ-ZrPO4(OH)1-x(RCOO)x(dmso) have been prepared by covalent anchoring of five different chiral carboxylic acids to the sheets of λ-zirconium phosphate (λ-ZrP), namely (S)-(+)-Mandelic acid (MAN), (R)-(−)-α-Methoxyphenylacetic acid (MPA), (S)-(+)-Ibuprofen (IBU), (S)-(+)-6-Methoxy-α-methyl-2-naphthaleneacetic acid (MMN) and (−)-Menthyloxyacetic acid (MEN). The synthesized materials were characterized by X-ray diffractometry, FT-IR spectrophotometry, elemental and thermogravimetric analyses and polarimetry. The X-ray diffraction patterns revealed that the interlayer separations increase from 1.02 nm (pure λ-ZrP) to 1.31–2.31 nm as a result of the incorporation of the mentioned acids within the interlayer gallery. The polarimetry measurements showed that the synthesized materials have appreciable optical activities suggesting they could have protentional applications in the field of chiral separation techniques.

Screening Approach for Identifying Cocrystal Types and Resolution Opportunities in Complex Chiral Multicomponent Systems

Cocrystallization of racemic-compound-forming chiral molecules can result in conglomerate cocrystals or diastereomerically related cocrystals, which enable the application of chiral separation techniques such as preferential crystallization and classic resolution. Here, a systematic method to identify the types and phase diagrams of cocrystals formed by chiral target compounds and candidate coformers in a particular solvent system is presented, which allows the design of suitable chiral resolution processes. The method is based on saturation temperature measurements of specific solution compositions containing both enantiomers of chiral molecules and a coformer. This method is applied to analyze three different systems. For racemic phenylalanine (Phe) in water/ethanol mixtures one of the enantiomers selectively cocrystallizes with the opposite enantiomer of valine (Val), forming the more stable diastereomerically related cocrystal. The racemic compound ibuprofen crystallizes with the nonchiral coformer 1,2-bis(4-pyridyl)ethane (BPN) as racemic compound cocrystals. More interestingly, when it is combined with trans-1-(2-pyridyl)-2-(4-pyridyl)ethylene (BPE), the racemic compound ibuprofen cocrystallizes as a conglomerate, which in principle enables the application of preferential crystallization of this racemic compound. The systematic method shows the benefit of using pseudo-binary phase diagrams. Such pseudo-binary phase diagrams depict the saturation temperature on a very specific route through the quaternary phase diagram, allowing the identification of various cocrystal types as well as the corresponding cocrystallization conditions. The systematic method can be used to identify a suitable solid phase for chiral separation, and the obtained phase diagram information enables the performance of a crystallization-mediated chiral resolution process design. Such a guideline for a chiral resolution process design has never been reported for conglomerate cocrystal systems such as IBU:BPE, presented in this study.

Enantiomeric Separation of Azole Antifungal Compounds using Chromatographic and Electrophoretic Techniques: A Mini Review

The separation of azole antifungal chiral compounds has gaining much interests since 1980’s as they possess different biological, pharmaceutical, and pharmacokinetic behaviors at various toxicity levels. Numerous studies have looked into this subject matter, and therefore, this review presents an overview on different chiral separation techniques that have been developed such as liquid chromatography (LC), gas chromatography (GC), supercritical fluid chromatography (SFC), and capillary electrophoresis (CE). This comprehensive review also discusses the mechanisms of the developed analytical methods such as preparation techniques and optimization parameters towards the enantioseparation performances as well as their advantages and drawbacks. Lastly, the conclusion and future directions of the chiral separation, particularly for azole compounds are provided accordingly.

Recent advances of separation and analysis of chiral compounds by capillary electrophoresis

As the enantiomers of chiral compounds often exhibit different physiological activities, the establishment of effective separation and analytical methods for chiral compounds is of great significance. As a widely used chiral separation and detection technique, capillary electrophoresis (CE) offers the advantages of high separation efficiency, high speed, small sample consumption and flexible separation mode. In this review, research progresses in chiral separation by CE for the period 2017 to 2019 are summarized, and future developments are prospected.

Cyclofructans as Chiral Selectors: An Overview.

Cyclofructans are cyclic oligosaccharides made of β-2,1-linked fructofuranose units. They have been utilized as chiral selectors, usually after derivatization, with high-performance liquid chromatography (HPLC), gas chromatography (GC), capillary electrophoresis (CE), and supercritical fluid chromatography (SFC). The focus herein will be directed to their development and applications as chiral selectors in various chiral separation techniques. Discussion of their use in hydrophilic liquid interaction chromatography (HILIC) will be limited. Their use in liquid chromatography, especially their improvements with the use of superficially porous particles (SPPs) will be emphasized. Method parameters and future directions are also discussed.

High-Throughput LC-MS/MS Method for Chiral Amino Acid Analysis Without Derivatization.

D-Amino acids have recently attracted much attention in various research fields including medical, clinical, and food industry due to their important biological functions that differ from L-amino acid. Most chiral amino acid separation techniques require complicated derivatization procedures in order to achieve the desirable chromatographic behavior and detectability. This chapter describes a highly sensitive analytical method for the enantioseparation of chiral amino acids without any derivatization process using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The method allows the simultaneous analysis of 18 D-amino acids with high sensitivity and reproducibility. Additionally, this chapter also focuses on the application of the method to real samples for the quantification of targeted amino acids.

Combining enzymatic hydrolysis with magnetic nanoparticles for resolution of chiral substances

At present, the separation of racemates is still a great challenge, since they are enantiomers, having similar chemical structures and physical properties. Here, we proposed a new strategy for efficiently separating chiral compounds, i.e., stereoselectively hydrolyzing the individual enantiomer of racemates via enzyme, followed by a simple magnetic separation using Fe3O4 nanoparticles, may implement a complete resolution of racemates. In this work, d-aminoacylase was selected as enzyme, and phenylalanine and methionine as chiral models. The use of d-aminoacylase can hydrolyze d-enantiomer, while l-enantiomer cannot. Magnetic nanoparticles of Fe3O4@SiO2-NH2-COOH composites were designed and synthesized for the amidation reaction with the hydrolyzed d-amino acids from the mixture. With the help of a magnetic field, d-form amino acids can be readily separated at a large scale and the resultant resolving performances indicated that this method exhibited a higher selectivity, compared with common chiral separation techniques. The enantiomeric excess (e.e.) values are 75% and 98% for racemic N-acyl-phenylalanine and methionine, respectively, which solely rely on the hydrolysis capability of used enzymes. This result suggests that our method is feasible, simple, easy to scale-up and high-efficient for the chiral resolution. These findings demonstrate that this strategy could provide new opportunities for improving enantioseparation performances for a host of enantio- and bio-substance with both high-yield and high-selectivity.

Interinstrumental Transfer of a Chiral Capillary Electrophoretic Method: The Use of Robustness Test Information to Overcome Differences in Detector and Data-Handling Specifications

Capillary electrophoresis has been widely used as chiral separation technique, applying chiral selectors that are added to the background electrolyte. The advantages of capillary electrophoresis as separation technique are its flexibility, low cost, and high separation efficiency. This study is part of a research project, where guidelines are defined to facilitate interinstrumental method transfer of capillary electrophoretic methods, which is one of the major drawbacks of capillary electrophoresis. Another drawback is the lower sensitivity compared to liquid chromatographic methods. Improving and maintaining the sensitivity are the reason why focus should be put on the interinstrumental differences between detector settings. The aim of this study was to determine when adaption of the detector settings during interinstrumental method transfer was needed. The chiral separations of two betablockers were selected as case studies. The influence of detector parameters, such as data acquisition rate, bandwidth, and filter, on sensitivity responses, such as peak area, height, and width, was evaluated by means of robustness tests performed on two capillary electrophoresis instruments. The statistically significant parameters were identified and non-significance intervals determined. To maintain or optimise the obtained sensitivity, the information gathered from the robustness test was further incorporated in guidelines developed to facilitate interinstrumental analytical method transfer of capillary electrophoretic methods.

Membrane Bioreactors for Pharmaceutical Applications: Optically Pure Enantiomers Production

In biological systems, recognition at molecular level is governed by chiral interactions. Therefore, optical isomers have very different effect in natural systems. For example, one can have beneficial effect while the other can be very harmful. For these reasons, chiral drugs nowadays are mainly admitted in the optically pure form. Given these requirements, it is clear why demand for chiral drugs has grown dramatically and the singleenantiomer drug segment has become an important part of the overall pharmaceutical market. As a consequence, the development of new chiral separation techniques is a very hot topic in both academic research and industrial innovation. Membrane bioreactors have proven their feasibility in the production of optically pure enantiomers by combining enantiospecific biochemical reactions with mass transport through membranes. The principles and the applications of enantioselective membrane bioreactors in kinetic resolution for pharmaceutical applications will be discussed. Various membrane bioreactors configurations and operation mode will be illustrated. The type of enzymes utilized to produce chiral drugs or their intermediates will be also reported. Multistep syntheses, conducted in sequential reactions catalysed by spatially aligned biocatalysts, as promising technology for the synthesis of fine chemicals will be highlighted.

Chiral separation of new sulfonamide derivatives and evaluation of their enantioselective affinity for human carbonic anhydrase II by microscale thermophoresis and surface plasmon resonance

The aim of this study was to develop a method combining chiral separation and biophysical techniques to evaluate the enantioselective affinity of original sulfonamide derivatives towards their therapeutic target, the human carbonic anhydrase II (hACII). The first step consisted in the preparation of the enantiomers by chromatographic separation. The performances of HPLC and Supercritical Fluid Chromatography (SFC) were studied at the analytical scale by optimization of various experimental conditions using adsorbed polysaccharide chiral stationary phases (amylose AD-H and cellulose OD-H). Since SFC allowed obtaining higher enantioresolutions per time unit, it was selected for the semi-preparative scale and successfully used to isolate each enantiomer with a satisfactory enantiomeric purity (>98%).Secondly, microscale thermophoresis (MST) method and surface plasmon resonance (SPR) used as reference method were developed to measure potential enantioselective affinities of these enantiomers towards the hACII. The optimizations of both methods were performed using a reference compound, i.e. acetazolamide, which affinity for hCAII has previously been demonstrated. For all compounds, KD values obtained using MST and SPR were in good agreement, leading to similar affinity scales despite both approaches totally differ (labeling for MST versus immobilization of the protein for SPR). The equilibrium dissociation constants of our original compounds for the hCAII were in the range 100–1000nM and an enantioselectivity was observed using the MST and SPR methods for the diarylpyrazole 2. Finally, by comparing the MST and SPR techniques, MST appears especially adapted for further screening of a series of sulfonamide derivatives due to the lower time required to estimate a binding constant while consuming as little hCAII as SPR.

Development of a liquid chromatography-tandem mass spectrometry method for quantitative analysis of trace d-amino acids

d-Amino acids have recently attracted much attention in various research fields including medical, clinical and food industry due to their important biological functions that differ from l-amino acid. Most chiral amino acid separation techniques require complicated derivatization procedures in order to achieve the desirable chromatographic behavior and detectability. Thus, the aim of this research is to develop a highly sensitive analytical method for the enantioseparation of chiral amino acids without any derivatization process using liquid chromatography-tandem mass spectrometry (LC-MS/MS). By optimizing MS/MS parameters, we established a quantification method that allowed the simultaneous analysis of 18 d-amino acids with high sensitivity and reproducibility. Additionally, we applied the method to food sample (vinegar) for the validation, and successfully quantified trace levels of d-amino acids in samples. These results demonstrated the applicability and feasibility of the LC-MS/MS method as a novel, effective tool for d-amino acid measurement in various biological samples.

Enantioseparation of phenylsuccinic acid enantiomers based on aqueous two-phase system with ethanol/ammonium sulfate: phase diagrams optimization and partitioning experiments

In the study herein we focused on enantioseparation of phenylsuccinic acid (PSA) enantiomers using an aqueous two-phase system (ATPS) composed of ethanol and ammonium sulfate. Based on phase-forming points and salting-out points, a modified phase diagram of ethanol/ammonium sulfate system with three distinct phase areas, including homogeneous area, ATPS area and salting-out area, has been constructed experimentally. The ATPS optimization and evaluation of chiral separation efficiency for PSA enantiomers have been investigated thoroughly. A hydrophilic chiral selector, hydroxypropyl-β-cyclodextrin (HP-β-CD), which has good chiral recognition ability for R-PSA, was added into bottom phase. The chiral separation efficiencies, namely distribution coefficients and separation factor, were affected by various process variables, including phase compositions of ATPS, concentrations of HP-β-CD and PSA enantiomers, pH value and operational temperature. Under the optimum conditions, separation factor could reach 1.42 after just one step extraction. This environmentally benign chiral separation technique opens up new possibilities for preparative separation of other racemic compounds at a large-scale.