Polar Organic Mode Research Articles

Ultrafast separation of fluorinated and desfluorinated pharmaceuticals using highly efficient and selective chiral selectors bonded to superficially porous particles

The separation of fluorinated active pharmaceutical ingredients (APIs) from their desfluoro analogs is a challenging analytical task due to their structural similarity. In this work, fluorine containing APIs and their corresponding desfluorinated impurities were separated on five new 2.7μm superficially porous particles (SPPs) functionalized with bonded chiral selectors. The unique shape selectivity of bonded macrocyclic glycopeptides and oligosaccharides was utilized to separate seven pairs of fluoro/desfluoro APIs resulting in some unprecedented selectivity values. For example, SPP bonded isopropyl cyclofructan 6 yielded a selectivity of 2.73 for voriconazole and desfluoro voriconazole. Further, the SPP based columns allowed for rapid separations ranging from 9 to 55s with very high efficiencies ranging from 45,000 to 70,000plates/m (at high flow rates) in both reversed phase and polar organic modes. Chromatographic separation and detection by HPLC-ESI-MS was demonstrated using ezetimibe and voriconazole and their desfluorinated impurities. Among the tested phases, SPP hydroxypropyl-β-cyclodextrin separated the most fluorinated and desfluorinated analogs with baseline resolution.

High efficiency, narrow particle size distribution, sub-2 μm based macrocyclic glycopeptide chiral stationary phases in HPLC and SFC

State of the art chiral chromatography still employs 3–5 μm bonded or immobilized chiral selectors in 10–25 cm columns. With the availability of 1.9 μm narrow particle size distribution (NPSD) silica, it is now possible to make ever shorter, high efficiency columns practical for sub-minute chiral separations. Three macrocyclic glycopeptides (teicoplanin, teicoplanin aglycone, and vancomycin) were bonded onto 1.9 μm NPSD particles. Such packed columns had ∼80% lower backpressure as compared to polydisperse (PD) 1.7 μm silica materials when using the same mobile phase. The decreased backpressure allowed for diminution of frictional heating and allowed for the use of the 1.9 μm NPSD particle based columns at high flow rates. The 1.9 μm NPSD particle based columns showed up to 190,000 plates m−1 for chiral molecules and 210,000 plates m−1 for achiral probes. Representative enantiomeric separations are shown for wide classes of compounds, including different types of amino acids, β-blockers, and pharmaceutically important heterocyclic compounds such as oxazolidinones. Applications in three liquid chromatography modes, namely, reversed phase, polar organic mode and normal phase chiral separations were shown with resolution values ranging from 1.5 to 5.7. Additionally, the same columns were used with supercritical fluid chromatography (SFC) for ultrafast separations.

Determination of the Enantiomerization Barrier of the Residual Enantiomers of C3 -Symmetric Tris[3-(1-Methyl-2-Alkyl)Indolyl]Phosphane Oxides: Case Study of a Multitasking HPLC Investigation Based on an Immobilized Polysaccharide Stationary Phase.

The residual enantiomers of three tris-(3-indolyl)-phosphane oxides bearing different alkyl groups (methyl, ethyl or i-propyl) in position 2 of the indole rings constituting the blades were separated on the immobilized type Chiralpak IC column in polar organic and reversed-phase modes. The good enantioselectivity and versatility of the IC CSP allowed easy isolation of the enantiomerically highly enriched samples suitable for configurational stability studies. The enantiomerization barriers of residual phosphane oxides were evaluated both by off-column techniques (CD signal and enantiomeric purity decay kinetics) and by dynamic enantioselective high-performance liquid chromatography (HPLC).

High performance liquid chromatographic enantioseparation development and analytical method characterization of the carboxylate ester of evacetrapib using an immobilized chiral stationary phase with a non-conventional eluent system

Using multiple HPLC chromatographic modes and various chiral columns in the context of an automated screening system, a potential separation was initially identified for the methyl ester of evacetrapib and its stereoisomers using an immobilized polysaccharide-based HPLC column. The bonded nature of this column, the Chiralpak® IC, allows for enhanced separation development with a diverse solvent range not amenable to standard coated chiral stationary phases. The ternary eluent system ultimately identified provided isomer resolutions not obtainable via the more established hexane/alcohol or polar organic chromatographic modes. A systematic separation development process is described, first for the resolution of the isomers, and later incorporating five potential impurities. A robust separation system was eventually developed that effectively resolves all compounds within a reasonable analysis time.

Separation Of Methionine Enantiomers By Using Teicoplanin And Cyclofructan Columns

Abstract Methionine is a naturally occurring amino acid. Its enantiomeric separation by using high performance liquid chromatography on various types of chiral stationary phases was studied. The effect of mobile phase composition on enantioselectivity and retention was considered. The separation of the enantiomers was attained in different separation modes – reversed phase mode for the macrocyclic antibiotic chiral stationary phases (teicoplanin, teicoplanin aglycone), normal phase and polar organic phase modes for the isopropyl carbamate cyclofructan 6 chiral stationary phase. It was shown that the hydrogen bonding, dipole interactions, steric effects between methionine molecules and stationary phases play an important role in the separation of enantiomers.

Preparation and Evaluation of HPLC Chiral Stationary Phases Based on Cationic/Basic Derivatives of Cyclofructan 6

The cyclofructan 6 (CF6) macrocyclic-oligosaccharide was derivatized with five different substituents able to bear positive charges: propyl imidazole, methyl benzimidazole, dimethyl aminopropyl, pyridine, and dimethyl aminophenyl. The derivatized cyclofructans were reacted with triethoxysilyl-propylisocyanate as a linker to bond them to 5 µm spherical silica gel particles and then used to prepare liquid chromatography columns. The bonded silica particles were analyzed to establish the bonding densities. A set of 34 chiral compounds including acids, neutral compounds, and bases was tested with nine different mobile phase compositions including two reverse phase (RP) acetonitrile/pH 4 buffer, three polar organic (PO) acetonitrile/methanol, and four normal phase (NP) heptane/ethanol mobile phases. No compounds could be separated in the RP mode. Eight compounds only could be enantioseparated in the PO mode and 21 compounds in the NP mode. The most effective chiral stationary phase was the propyl imidazole derivatized CF6 phase, provided no more than six imidazole substituents and two linkers are attached per CF6 unit.

Determination of methionine enantiomers by HPLC on the cyclofructan chiral stationary phase

The enantiomeric separation of methionine by high performance liquid chromatography on various cyclofructan type chiral stationary phases in the polar-organic separation mode was studied.

Preparation of β-Cyclodextrin Capillary Column for Enantioseparation and Determination of Nicardipine and Manidipine by Capillary Electrochromatography

The separation and determination methods of calcium antagonists manidipine hydrochloride and nicardipine hydrochloride enantiomers were developed by open-tubular capillary electrochromatography( CEC),respectively. A new capillary column with p-nitrophenylcarbamation β-cyclodextrin was prepared by sol-gel method. The sol-gel modified with cyclodextrin derivative was characterized by infrared spectroscopy( IR) and scanning electron microscopy( SEM). In polar organic mode,the effect of the volume fraction and concentration of added triethylamine and glacial acetic acid,the content of organic solvents,the operating voltage,and temperature on the resolution( RS) were investigated. The optimized conditions were at 20 ℃,detection wavelength at 236 nm,and injection pressure at 3. 448 k Pa × 3 s. The compositions of mobile phases were V( Me OH) ∶ V( ACN) ∶ V( TEA) ∶ V( HOAc) = 57 ∶ 43 ∶ 0. 05 ∶ 0. 07 for manidipine hydrochloride and V( Me OH) ∶ V( ACN) ∶ V( TEA) ∶ V( HOAc) = 55 ∶ 45 ∶ 0. 05 ∶ 0. 08 for nicardipine hydrochloride. The resolution( Rs) of manidipine hydrochloride was 1. 39 at 25 k V. The resolution( Rs) of nicardipine hydrochloride was1. 30 at 20 k V. The RSDs of retention time and peak area were less than 1. 2% and 5. 6%( n = 5) for the drugs,respectively. It was indicated that new column has stable electrochromatography performance. The single run time were less than 5 min for manidipine hydrochloride and less than 7 min for nicardipine hydrochloride. Good linearity were observed in the concentration range of 5. 2 ~ 125. 0 mg / L with correlation coefficients 0. 9969 or more for both drug enantiomers. The limit of detection values were 1. 8 mg / L for manidipine and 2. 3 mg / L for nicardipine,respectively. The nitrophenyl carbamated β-cyclodextrin phase( NCDP) column were preliminarily used for determinations of manidipine hydrochloride and nicardipine hydrochloride tablet and capsule.

Enantiomeric separations of ruthenium (II) polypyridyl complexes using HPLC with cyclofructan chiral stationary phases.

The enantiomeric separation of 21 ruthenium (II) polypyridyl complexes was achieved with a novel class of cyclofructan-based chiral stationary phases (CSPs) in the polar organic mode. Aromatic derivatives on the chiral selectors proved to be essential for enantioselectivity. The R-napthylethyl carbamate functionalized cyclofructan 6 (LARIHC CF6-RN) column proved to be the most effective overall, while the dimethylphenyl carbamate cyclofructan 7 (LARIHC CF7-DMP) showed complementary selectivity. A combination of acid and base additives was necessary for optimal separations. The retention factor vs. acetonitrile/methanol ratio plot showed a U-shaped retention curve, indicating that different interactions take place at different polar organic solvent compositions. The separation results indicated that π-π interactions, steric effects, and hydrogen bonding contribute to the enantiomeric separation of ruthenium (II) polypyridyl complexes with cyclofructan chiral stationary phases in the polar organic mode.

Erratum to: Chiral Separation of 4-Iminoflavan Derivatives on Several Polysaccharide-Based Chiral Stationary Phases by HPLC

The HPLC enantiomeric separation of seven 4-iminoflavans was successfully accomplished in the normal phase mode using six polysaccharide-based chiral stationary phases namely, Chiralcel®OD-H, Chiralcel®OD, Chiralcel®OJ, Chiralpak®AD, Chiralpak®IA and Chiralpak®IB under normal and polar organic phase modes. The resolution depended on nature and concentration of alcoholic modifier. The results demonstrate clearly that the chromatographic system based on the coated and immobilized type Chiralpak®IB and Chiralcel®OD-H CSPs provide a powerful analytical tool for enantiomeric separation of all the 4-iminoflavans used in this study.

High‐performance liquid chromatography enantioseparation of polyhalogenated 4,4′‐bipyridines on polysaccharide‐based chiral stationary phases under multimodal elution

An investigation on the high-performance liquid chromatography enantioseparation of 12 polyhalogenated 4,4'-bipyridines on polysaccharide-based chiral stationary phases is described. The overall study was directed toward the generation of efficient separations in order to obtain pure atropisomers that will serve as ligands for building homochiral metal organic frameworks. Four coated columns--namely, Lux Cellulose-1, Lux Cellulose-2, Lux Cellulose-4, and Lux Amylose-2--and two immobilized columns--namely, Chiralpak IC and IA--were used under normal, polar organic, and reversed-phase elution modes. Moreover, Chiralcel OJ was considered under normal-phase and polar organic conditions. The effect of the chiral selector and mobile phase composition on the enantioseparation, the enantiomer elution order and the beneficial effect of nonstandard solvents were studied. The effect of water in the mobile phase on the enantioselectivity and retention was investigated and retention profiles typical of hydrophilic interaction liquid chromatography were observed. Interesting phenomena of solvent-induced enantiomer elution order reversal occurred under normal-phase mode. All the considered 4,4'-bipyridines were enantioseparated at the multimilligram level.

Enantioseparation and Determination of Propranolol in Human Plasma on a New Derivatized β-Cyclodextrin-Bonded Phase by HPLC

A 6-azido-β-cyclodextrin was synthesized and derivatized with p-nitrophenyl isocyanate as chiral ligand. Then the ligand was chemically bonded to mesoporous SBA-15 via a ‘Click Chemistry’ reaction of the azido group with alkynyl group. A new p-nitrophenylcarbamoylated β-cyclodextrin bonded SBA-15 chiral stationary phase (NPCSP) for HPLC was thus obtained. The new stationary phase was first used for enantioseparation of propranolol in human plasma under the polar organic solvent mode. The effects of methanol content, concentration of glacial acetic acid/triethylamine in mobile phase and the temperature on the enantioseparation were studied in detail. The optimized chromatographic conditions were as follows: mobile phase was acetonitrile/methanol/glacial acetic acid/triethylamine (90/10/1.25/2.25, V/V) at 288 K, with a flow rate of 0.5 mL min−1, an injection volume of 20 μL and detection wavelength at 290 nm. The resolution was 2.04 with a short run time (< 15 min) under the optimal conditions. The content of propranolol in plasma was quantitatively measured by HPLC-MS under selected ion monitoring mode ([M + H]+ m/z 260.10) with hydrochlorothiazide as an internal standard. The good linear relationship was observed in the range of 2.5−50 μg L−1. The detection limits were 1 μg L−1 according to S/N = 3 for both enantiomers. The experimental results showed that the chiral stationary phase exhibited excellent chiral separation ability to propranolol. The analytical method for propranolol enantiomers was sensitive, accurate, simple and fast, which was can be for the determination of propranolol in plasma and related pharmacokinetic studies.

Novel chiral stationary phases based on peptoid combining a quinine/quinidine moiety through a C9‐position carbamate group

By connecting a quinine or quinidine moiety to the peptoid chain through the C9-position carbamate group, we synthesized two new chiral selectors. After immobilizing them onto 3-mercaptopropyl-modified silica gel, two novel chiral stationary phases were prepared. With neutral, acid, and basic chiral compounds as analytes, we evaluated these two stationary phases and compared their chromatographic performance with chiral columns based on quinine tert-butyl carbamate and the previous peptoid. From the resolution of neutral and basic analytes under normal-phase mode, it was found that the new stationary phases exhibited much better enantioselectivity than the quinine tert-butyl carbamate column; the peptoid moiety played an important role in enantiorecognition, which controlled the elution orders of enantiomers; the assisting role of the cinchona alkaloid moieties was observed in some separations. Under acid polar organic phase mode, it was proved that cinchona alkaloid moieties introduced excellent enantiorecognitions for chiral acid compounds; in some separations, the peptoid moiety affected enantioseparations as well. Overall, chiral moieties with specific enantioselectivity were demonstrated to improve the performance of peptoid chiral stationary phase efficiently.

Enantiomeric Separation of Bicyclo[2.2.2]octane‐Based 2‐Amino‐3‐Carboxylic Acids on Macrocyclic Glycopeptide Chiral Stationary Phases

Direct high-performance liquid chromatographic (HPLC) separation of four bicyclo[2.2.2]octane based 2-amino-3-carboxylic acid enantiomers were developed on chiral stationary phases (CSPs) containing different macrocyclic glycopeptide antibiotic selectors. The analyses were performed under reversed-phase, polar organic and polar ionic mode on macrocyclic-glycopeptide-based Chirobiotic T, T2, TAG, and R columns. The effects of the mobile phase composition including the acid and base modifier, the structure of the analytes, and the temperature on the separations were investigated. Experiments were achieved at constant mobile phase compositions on different stationary phases in the temperature range 5-40°C. Thermodynamic parameters were calculated from plots of ln k or ln α versus 1/T. It was recognized that the enantioseparations in reversed-phase and polar organic mode were enthalpically driven, but under polar-ionic conditions entropically driven enantioseparation was observed as well. Baseline separation and determination of elution sequence were achieved in all cases.

Novel carbamoyl type quinine and quinidine based chiral anion exchangers implementing alkyne–azide cycloaddition immobilization chemistry

The synthesis and chromatographic evaluation of a series of new Cinchona derived chiral weak anion exchangers is presented. Huisgen Cu(I) mediated alkyne–azide cycloaddition, so-called click chemistry, was used as an immobilization strategy. In this way it was possible to immobilize about 90% of offered selector via 1,2,3-triazole linker, which displays a more efficient way of binding the selector to modified silica compared to common radical mediated thiol-ene addition. Problems associated with potential radical scavenging properties of chiral selectors thereby could be circumvented. The evaluation of the synthesized chiral stationary phases regarding chromatographic behavior was carried out using polar organic mode mobile phase composition and a set of representative chiral organic acids. Different loading densities revealed an optimum selector density of about 310μmol/g chiral stationary phase with respect to resolution and selectivity. A decrease of performance was observed for higher loading, indicating mutual spatial influence of selector units leading to sterical hindrance. In addition, we observed that the effect of free azide groups on retention is negligible and the overall chromatographic behavior is comparable to other Cinchona derived chiral stationary phases.

Preparation and Evaluation of a Novel bis(β-cyclodextrin)-bonded SBA-15 Chiral Stationary Phase for HPLC

由于桥联双β-环糊精的协同包结作用和独特的多重识别功能已被广泛地用作人工模拟酶,本文尝试用桥联双β-环糊精作固定相配体,开发其手性分离功能.采用3-异氰酸丙基三乙氧基硅烷偶联剂连续反应法,首先将(6-氧-对甲苯磺酰)-β-环糊精键合到有序介孔SBA-15硅胶表面,然后与(6-乙二胺-6-去氧)-β-环糊精反应,制备一种新型的乙二胺桥联双β-环糊精键合SBA-15液相色谱手性固定相(BCDSP).采用质谱、红外光谱、元素分析、热重分析和透射电镜等进行结构表征.在极性有机溶剂模式下,评价了新固定相的基本色谱性能,并用于β-受体阻滞剂对映体的拆分.考察了流动相中有机溶剂、三乙胺、冰醋酸改性剂用量和柱温对手性分离的影响.基于优化的色谱条件,采用双β-环糊精键合固定相成功地拆分了常用的14种β-受体阻滞剂药物对映体,其中普萘洛尔的分离度可达到2.18,卡维地洛的分离度可达到2.01,分析时间一般为10~20 min,取得了较高的分离效率.为比较研究,还制备了一种单β-环糊精固定相(CDSP),结果发现双β-环糊精键合相可拆分14种β-受体阻滞剂,而在优化的条件下单β-环糊精键合相仅能部分拆分4种β-受体阻滞剂,且前者的分离度明显优于后者.基于实验数据对双β-环糊精和单β-环糊精固定相的分离机理的异同点展开讨论.一方面包结作用和氢键作用在桥联双β-环糊精键合相的手性分离中占重要地位,与常见的单β-环糊精固定相相似;另一方面桥联双β-环糊精固定相拥有自身的特点,两个β-环糊精腔体存在协同作用,相应于它的模拟酶功能,协同作用不仅扩大了空间识别域,还提高了立体选择性,基于双腔体的包结作用和乙二胺桥键的氢键作用等增强了固定相的手性识别能力,加上有序的SBA-15能加快传质,使得BCDSP具有较好的手性色谱性能,拥有更广泛的拆分对象、更高的分�

Enantioseparation and determination of atenolol enantiomers in tables on a β-cyclodextrin-based chiral stationary phase by high performance liquid chromatography

A novel dinitrophenyl ether β-cyclodextrin-bonded chiral stationary phase (NESP) for HPLC was prepared with ordered mesoporous SBA-15 as matrix. The fast enantioseparation of atenolol enantiomers on the new stationary phase was achieved through the optimization of mobile phase composition, column temperature and other factors in polar organic solvent mode. The optimized composition of mobile phase was acetonitrile/methanol/glacial acetic acid/triethylamine (90:10:2.5:3.0, v/v/v/v) at the flow rate of 0.5 mL/min. The column tem- perature was set at 20 °C. The detection wavelength was 275 nm. The resolution of atenolol enantiomers was 1.73 with a rather short analysis time, about 20 min, under the above conditions. The atenolol was extracted with methanol from the tablets and analyzed by direct injection for HPLC. A new quantitative method of atenolol enantiomers in tablets was established after the condition optimization. The good linear relationships for two atenolol enantiomers were observed in the range of 2.5-100 mg/L with r of 0. 999 2 and 0. 998 9, respectively. The recoveries of atenolol enantiomers in tablet samples were 94.60%-97.24%. The relative standard deviations (RSDs) of this method were not greater than 0.92% for intra-day and 1.86% for inter-day, respectively. The detection limits (S/N = 3) of concentrations were less than 0.2 mg/L for both enantiomers. The established method is simple and of high selectivity, high recovery and low cost for enantiomer analysis by using homemade cyclodextrin-based chiral column. It has a good application prospect for the fast quality control and the pharmacokinetics study of chiral drugs.

Enantiomeric Separations of Chiral Sulfonic and Phosphoric Acids with Barium-Doped Cyclofructan Selectors via an Ion Interaction Mechanism

New cyclofructan-6 (CF6)-based chiral stationary phases (CSPs) bind barium cations. As a result, the barium-complexed CSPs exhibit enantioselectivity toward 16 chiral phosphoric and sulfonic acids in the polar organic mode (e.g., methanol or ethanol mobile phase containing a barium salt additive). Retention is predominantly governed by a strong ionic interaction between the analyte and the complexed barium cation as well as hydrogen bonding with the cyclofructan macrocycle. The log k versus log [X], where [X] = the concentration of the barium counteranion, plots for LARIHC-CF6-P were linear with negative slopes demonstrating typical anion exchange behavior. The nature of the barium counteranion also was investigated (acetate, methanesulfonate, trifluoroacetate, and perchlorate), and the apparent elution strength was found to be acetate > methanesulfonate > trifluoroacetate > perchlorate. A theory based upon a double layer model was proposed wherein kosmotropic anions are selectively adsorbed to the cyclofructan macrocycle and attenuate the effect of the barium cation. van't Hoff studies for two analytes were conducted on the LARIHC-CF6-P for three of the barium salts (acetate, trifluoroacetate, and perchlorate), and the thermodynamic parameters governing retention and enantioselectivity are discussed. Interestingly, for the entropically driven separations, enantiomeric selectivity can increase at higher temperatures, even with decreasing retention.

Enantioselective separation of unusual amino acids by high performance liquid chromatography

Unusual amino acids, i.e. amino acids not encoded by DNA, play fundamental roles in many scientific fields. Since the single enantiomeric form can cause different and often serious response of organisms, chiral separations of unusual amino acids are irreplaceable tools in their study and their employment. Two types of chiral stationary phases, two teicoplanin-based and four polysaccharide-based columns, were used. Separation conditions of reversed phase mode, polar organic mode and hydrophilic interaction chromatography were evaluated and compared. All columns used exhibited significantly different enantioselectivities. Teicoplanin-based chiral stationary phases, especially Chirobiotic T column, were able to separate almost all enantiomers tested, with the exception of Z-D,L-4-F-phenylalanine ethyl ester. No partial enantioseparation of this analyte was obtained on teicoplanin-based chiral stationary phases, while baseline enantioresolution was achieved on polysaccharide-based columns. Change of elution order of L- and D-enantiomers was proved regarding the chiral stationary phase used.

Enantiomeric separations of illicit drugs and controlled substances using cyclofructan‐based (LARIHC) and cyclobond I 2000 RSP HPLC chiral stationary phases

Recently a novel class of chiral stationary phases (CSPs) based on cyclofructan (CF) has been developed. Cyclofructans are cyclic oligosaccharides that possess a crown ether core and pendent fructofuranose moieties. Herein, we evaluate the applicability of these novel CSPs for the enantiomeric separation of chiral illicit drugs and controlled substances directly without any derivatization. A set of 20 racemic compounds were used to evaluate these columns including 8 primary amines, 5 secondary amines, and 7 tertiary amines. Of the new cyclofructan-based LARIHC columns, 14 enantiomeric separations were obtained including 7 baseline and 7 partial separations. The LARIHC CF6-P column proved to be the most useful in separating illicit drugs and controlled substances accounting for 11 of the 14 optimized separations. The polar organic mode containing small amounts of methanol in acetonitrile was the most useful solvent system for the LARIHC CF6-P CSP. Furthermore, the LARIHC CF7-DMP CSP proved to be valuable for the separation of the tested chiral drugs resulting in four of the optimized enantiomeric separations, whereas the CF6-RN did not yield any optimum separations. The broad selectivity of the LARIHC CF7-DMP CSP is evident as it separated primary, secondary and tertiary amine containing chiral drugs. The compounds that were partially or un-separated using the cyclofructan based columns were screened with a Cyclobond I 2000 RSP column. This CSP provided three baseline and six partial separations.