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Carbamoylated derivatives of two antibiotics, namely, clindamycin phosphate (CLIP) and erythromycin (ERY) were successfully employed as co-precursors, in combination of zirconium tetrabutoxide as a precursor, to prepare chiral organic-zirconia hybrid monoliths (i.e., CLIP-ZHMs and ERY-ZHMs, respectively) via a single-step in-situ sol-gel approach in our previous works. Their superiority over chiral organic-zirconia/silica monoliths, prepared by post-modification approach, in terms of better enantioresolution and enhanced stability inspired us to prepare ZHMs based on an another antibiotic, azithromycin (i.e., AZI-ZHMs). Monolithic columns were employed for capillary electrochromatographic enantioseparation of acidic chiral drugs in mobile phases consisting of acetonitrile (ACN) and methanol (MeOH) as organic modifiers, and acetic acid (AcOH) and triethylamine (TEA) as electrolytes. The effects of composition of mobile phase and applied voltage on chiral separation were investigated by using ketoprofen as a representative analyte. Baseline resolutions were obtained for six acidic drugs in mobile phase consisting of 80/20 ( v / v ) ACN/MeOH with 300 mM AcOH and 10 mM TEA at a 10 kV applied voltage and 25 °C capillary temperature. The relative standard deviations for resolution values regarding column to column and batch to batch repeatability were less than 2.5% (for n = 3) under optimized conditions, indicating satisfactory stability of the columns and reproducibility of the column preparation process.

Rifampicin, a member of rifamycin sub-class of antibiotics which belongs to the naphthalenic ansamycin class of antibiotics, has a characteristic ansa structure, i.e., a ring structure or chromophore spanned by an aliphatic chain. The present work was designed to evaluate its potential as a chiral selector (CS) as its structure consisting of nine stereogenic centers, an aromatic moiety and several functional groups (i.e., one imine, one amide, one acetoxy residue, two aliphatic hydroxyl and three phenolic hydroxyl groups) was expected to instigate multiple enantioselective interactions, namely, hydrogen bonding and inclusion complexation with chiral analytes, and therefore resulting in efficient enantioseparations. Systematic experiments were performed to investigate the effects of concentration of CS, composition of background electrolyte (BGE) and applied voltage on chiral separation. Enantiomers of propranolol and metoprolol were baseline resolved using a BGE consisting of 20 mM CS and 50/50 ( v / v ) iso -propanol/phosphate buffer (100 mM, pH 7.0) whereas for enantiomers of sertraline, a BGE consisting of 23 mM CS and 40/60 ( v / v ) iso -propanol/phosphate buffer (100 mM, pH 7.0) resulted in baseline resolutions.

A tert-butylcarbamoylquinine ( tBuCQUI)-incorporated silica hybrid monolithic (QUI–SHM) capillary was prepared by “one-pot” approach via simultaneous sol–gel reaction of tetramethoxysilane and vinyltrimethoxysilane and free radical polymerization of tBuCQUI. A set of acidic chiral drugs were enantioseparated on QUI–SHM in mobile phases consisting of acetonitrile (ACN), methanol (MeOH), acetic acid (AcOH), and triethylamine (TEA). Influences of ACN/MeOH ratio, AcOH/TEA ratio, and their concentrations on enantioseparation were investigated. Baseline separations of enantiomers of all the compounds investigated were achieved with a mobile phase of 60:38 (v/v) ACN/MeOH containing 350 mM AcOH and 30 mM TEA under the applied voltage of −10 kV at 25°C.

An organic-inorganic hybrid monolithic column was prepared within the confines of a capillary via a single-step in situ sol-gel approach using zirconium tetrabutoxide as a precursor to compose the inorganic backbone and 3-triethoxysilylpropyl carbamoylated derivative of erythromycin (TEOSPC-ERY) as a co-precursor to introduce the organic chiral selector moiety in the zirconia backbone. The resulting carbamoylated ERY-zirconia hybrid monolith (ERY-ZHM) showed homogeneous morphology with well-defined through pores and was tightly connected with the inner wall of the capillary. The column was employed for capillary electrochromatographic enantioseparation of six basic chiral drugs in mobile phases (MPs) consisting of acetonitrile (ACN) and triethylammonium acetate (TEAA) buffer. The effects of composition of MP and applied voltage on chiral separation were investigated by using propranolol as a representative analyte. The highest resolution (Rs=3.33) was obtained with a MP consisting of 10/90 (v/v) ACN/TEAA buffer (10mM, pH 7), 10 kV applied voltage and 25°C capillary temperature. The relative standard deviations for resolution values regarding run to run, day to day, column to column and batch to batch repeatability were 0.41%, 0.89%, 1.80% and 2.26% (for n=3), respectively, indicating satisfactory stability of columns and reproducibility of column preparation process.

Doxycycline (DOX) is one of the tetracycline class of antibiotics and has not been examined for its enantioseparation abilities previously. The purpose of the present work was to evaluate DOX as a chiral selector (CS) using capillary electrophoresis (CE) in nonaqueous mode. Systematic experiments were performed to investigate the effects of concentration of CS, compositions of organic solvents and background electrolytes, applied voltage on chiral separation of a set of acidic chiral compounds. Excellent resolutions of enantiomers of acidic chiral compounds were attained in a background electrolytes composed of 50:50 acetonitrile/methanol+214mM acetic acid+ 63mM triethylamine and 38mM DOX. The results show that DOX is a viable chiral selector in CE for the enantioseparation of the type of chiral compounds investigated.

The penicillin sub-class of β-lactam antibiotics has not been examined for its enantiodiscriminating abilities in capillary electrophoresis (CE) until date. The present work was therefore designed to evaluate penicillin G potassium salt (PenG) as an ion-pair chiral selector (CS) using CE for its several attributes, namely, high solubility in water and lower alcohols, structure allowing multiple interactions with analytes and cost-effectiveness. Systematic experiments were performed to investigate the effect of composition of background electrolyte, applied voltage and capillary temperature on chiral separation. Baseline resolutions of enantiomers of five basic chiral drugs (namely, darifenacin, citalopram, sertraline, propranolol and metoprolol) were attained using a background electrolyte composed of water:methanol (90:10, v/v) and consisting of 10.7 or 16.1mM CS at 20°C using an applied voltage of 5kV.

E-mail: jhpark@ynu.ac.krReceived January 28, 2013, Accepted March 24, 2013Sulfated β-cyclodextrin (SCD)-coated zirconia monolith was used as the chiral stationary phase in capillaryelectrochromatography for enantiomeric separation of basic chiral compounds. SCD adsorbed on the zirconiasurface provided a stable chiral stationary phase in reversed-phase eluents. Retention, chiral selectivity andresolution of a set of six basic chiral compounds were measured in eluents of varying pH, composition ofmethanol and buffer. Optimum mobile phase condition for the separation of the compounds was found to bemethanol content of 30%, buffer concentration of 30 mM and pH of 4.0.Key Words : Chiral separation, Capillary electrochromatography, Zirconia monolith, Sulfated β-cyclodextrinIntroductionCapillary electrochromatography (CEC) on monolithiccolumns are becoming attractive alternative to particle-packed columns in HPLC and electrochromatography.

Recent years have witnessed several new trends in chiral separation, for example, the enantiorecognition ability of several new antibiotics has been explored using capillary electrophoresis (CE) prior to HPLC; antibiotics have been employed as chiral selectors (CSs) in a nonaqueous CE (NACE) mode; and several new detection techniques (namely, capacitively coupled contactless conductivity detection) have been used in combination with CE for quantification of enantiomers. On account of these emerging trends, this article aims to review the application of various classes of antibiotics for CE enantioseparation of pharmaceuticals. A detailed account of the basic factors affecting enantioseparation, certain limitations of antibiotics as CSs and strategies to mitigate them, and advantages of NACE while using antibiotics as CSs has also been presented. Copyright © 2013 John Wiley & Sons, Ltd.

Sulfonated cellulose tris(3,5-dimethylphenylcarbamate) (SCDMPC)-coated zirconia monolith (ZM) was used as the chiral stationary phase in capillary electrochromatography for separation of enantiomers of ten chiral compounds in acetonitrile (ACN)-phosphate buffer mixtures as the eluent. Influences of the ACN content, buffer concentration and pH on chiral separation have been investigated. Separation data on SCDMPC-ZM have been compared with those on CDMPC-ZM. Resolution factors were better on SCDMPC-ZM than CDMPC-ZM while retention factors were in general shorter on the former than the latter. Best chiral resolutions on SCDMPC-ZM were obtained with the eluent of 50% ACN containing 50 mM phosphate at pH around 4.

Porous zirconia monolith modified with clindamycin phosphate (CLIP-ZM) was used as chiral stationary phase (CSP) to separate a set of six acidic and basic chiral compounds in capillary electrochromatography (CEC). Resolutions and chiral selectivity factors of the chiral compounds were measured in ACN/MeOH mobile phases of varying compositions of MeOH and ammonium acetate as the electrolyte. In contrast to the CE separations where only chiral separations of acidic compounds were achieved enantiomers of both acidic and basic compounds were separated with acidic compounds better resolved than basic ones by CEC on the CLIP-ZM CSP. Best chiral separations of the chiral compounds were obtained with an eluent containing 10mM ammonium acetate and 35 vol.% methanol.

Separation of enantiomers is one of the most interesting and challenging tasks in the field of separation science. High-performance liquid chromatography (HPLC) and recently capillary electrochromatography (CEC) using chiral stationary phases (CSP) are two of the most useful techniques for both enantioseparation and determination of enantiomeric composition. Although silica has been extensively used as the base material for the stationary phases due to attractive properties including desirable mechanical strength, surface characteristics, catalytic inertness, and surface derivatization possibilities, it has some inherent shortcomings including the narrow range of pH and thermal stability. The zirconia-based stationary phases have shown excellent performance in the separation of wide range of compounds and have been considered as an alternative to silica-based stationary phases due to desirable chemical and physical properties such as resistance to thermal conditions, resistance to extreme pHs, and mechani...

Porous zirconia monolith (ZM) modified with cellulose 3,5-dimethylphenylcarbamate (CDMPC) was used as chiral stationary phaseto separate basic chiral compounds in capillary electrochromatography. The electroosmotic flow behavior of bare and CDMPC-modified zirconia monolithic (CDMPC-ZM) column was studied in ACN/phosphate buffer eluents of pH ranging from 2 to 12. The CDMPC-ZM column was evaluated by investigatingthe influences of pH, the type and composition of organic modifier of the eluent on enantioseparation. CEC separations at pH 9 provided the best resolutions for the analytes studied, which are better than those observed on CDMPC-modified silica monolithic columns under similar chromatographic conditions. No appreciable decline in retention and resolution factors after over 200 injections, and run-to-run and day-to-day repeatabilities of the column of less than 3% indicate the stability of the zirconia monolithic column in basic media.

Cellulose tris(3,5-dimethylphenylcarbamate) (CDMPC) is an excellent chiral selector for enantioseparation of a wide variety of chiral compounds. The monolithic chiral columns are becoming popular in liquid chromatography and capillary electrochromatography. In this work, we present the fast separation of chiral β-blockers on a CDMPC-modified zirconia monolithic column by capillary electrochromatography (CEC). The porous zirconia monolithic capillary column was prepared by using the sol-gel technology and then zirconia surface modified with CDMPC. The enantioseparations were performed in reversed-phase (RP) eluents of a phosphate solution (pH 4.4) modified with acetonitrile or alcohol. The enantioseparations of a set of eight chiral β-blockers were achieved in less than one minute. Influences of the applied voltage, column temperature, concentration of acetonitrile and the type of alcohol as the organic modifier in the mobile phase, and sample injection time on enantioseparation were investigated. CEC separations at the applied voltage of 10 kV and 15 °C in the ACN-modified mobile phase provided the best resolutions for the analytes studied. Run-to-run and day-to-day repeatabilities of the column in the RP-CEC separation were less than 1 and 2%, respectively.

A gradient reversed-phase high performance liquid chromatography (RP-HPLC) method was developed for separation and quantitation of impurities in pharmaceutical dosage form of ofloxacin tablets. The developed method was a stability indicating test method for estimation of related impurities generated during synthesis, formulation, and storage of ofloxacin tablets. Forced degradation studies were performed on ofloxacin tablets including acid hydrolysis (5.0 M hydrochloric acid), base hydrolysis (5.0 M sodium hydroxide), oxidation (30% hydrogen peroxide), heat (105°C) humidity degradation 25°C/92% RH/119 b & 40 min, and photolytic degradation (2600 Lux/119 h & 40 min). From the degradation study, the degradation was found between 0–15%. Limit of detection and limit of quantification were established in terms of percentage for all potential impurities. The recovery studies were conducted on finished dosage samples (tablets) for all potential impurities and the average percentage recovery was ranged from 90.8 ...

(-)-(18-Crown-6)-2,3,11,12-tetracarboxylic acid-bonded silica was used as the chiral stationary phase in capillary electrochromatography (CEC) for enantioseparation of some alpha-amino acids. Separation data in CEC were measured in mobile phases of varying pH, and composition of methanol and buffer, and compared with those in capillary liquid chromatography (CLC). In CEC better enantioseparation was generally obtained in the eluent of lower pH, higher buffer concentration and intermediate MeOH content, usually at the expense of analysis time. CEC showed generally better enantioselectivity and resolutions than CLC for the amino acids investigated.

chiral analytes in HPLC and recently in CEC. In this work we report enantio-separation of non-acidic chiral analytes on a quinine carbamate-immobilized zirconia (QNZ) in reversed-phase (RP) CEC. Influences of pH, composition of the buffer, acetonitrile content and the applied voltage on enantioseparation were examined. Enantiomers of the analytes investigated are well separated in acetonitrile/phosphate buffer mobile phases. Separation data on QNZ were compared to those on QN-bonded silica (QNS). Retention was longer but better enantioselectivity and resolution were obtained on QNZ than QNS.

A liquid chromatography–mass spectrometry (LC-MS) method was developed and validated for determining the levels of 4-methyl-2-cyano biphenyl and 4-bromomethyl-2-cyano biphenyl, which are key starting materials of an antihypertensive drug substance, irbesartan. An active pharmaceutical ingredient of irbesartan was synthesized by using these two starting materials for its therapeutic use. We have explicated the LC-MS method to separate and quantify these two compounds in irbesartan at nanogram levels. The method was capable of separating irbesartan and its starting materials, which were monitored for their absence in the finished product of irbesartan. The separation was carried out at 40°C on a 150- × 4.6-mm cyano column by using the mobile phase containing 60 volumes of water adjusted to pH 3.2 with formic acid and 40 volumes of acetonitrile. The detection wavelength was 220 nm. The MS involved an electrospray ionization (ESI) probe and ion-trap analyzer and was validated with respect to its spec...

(S)-N-(3,5-dinitrobenzoyl)leucine-N-phenyl-N-propylamine-bonded silica was used as a chiral stationary phase for separation of a set of racemic pi-acidic and pi-basic alpha-amino acid amides in electrolyteless ACN-water eluents by CEC in the RP and polar organic (PO) modes. The effect of the amount of water in the ACN-water eluent on chiral separation was examined. As water is added to ACN, retention was shortened but resolution and selectivity deteriorated severely. Retention, enantioselectivity, and resolution factors obtained in 100% ACN were compared with those in an n-hexane-isopropanol eluent with a small amount of water by normal phase (NP) CEC. Much shorter retention times with comparable enantioselectivities were observed with 100% ACN, demonstrating the advantage of separation on (S)-N-(DNB)leucine-N-phenyl-N-propylamine-bonded silica in PO-CEC over NP-CEC.

(S)-N-(3,5-Dinitrobenzoyl)leucine-N-phenyl-N-propylamide-bonded silica was used as a chiral stationary phase for separation of enantiomers of some racemic pi-acidic and pi-basic amino acid amides in normal phase capillary LC (CLC) and CEC. For generation of EOF in CEC, different amounts of water were added to n-hexane-isopropanol (IPA) eluents. Influences of added water and composition of polar modifier on retention, enantioselectivity and resolution were studied. Generally better separation was obtained in the eluents of lower water content. Shorter retention and better enantioselectivities were observed in the eluents of higher IPA content, whereas resolution became lower as IPA content was increased. Enantioselectivities were comparable but resolutions were much better in CEC than CLC.

Hexane waspurchased from EM Sciences (Gibbstown, HPLC-grademethanol was obtained from J.T. Baker (Phillipsburg, USA).Water was processed with an Elgastat UHQ water purifi-cation system (Bucks, UK). All the chemicals were of thebest quality available and used as received without anyfurther purification.

Spherical porous silica supports modified with titanium or zirconium alkoxides were prepared, and allyl groupswere chemically attached to the titanized or zirconi zed silica supports, and the product was cross-polymerizedwith a double bond containing cellulose derivative to yield new CSPs (chiral stationary phases). Magic anglespinning

Liquid chromatographic enantiomer separation of N‐phthaloyl (PHT) protected α‐amino acids on several coated and immobilized chiral stationary phases (CSPs) derived from polysaccharide derivatives was performed. The coated CSP of Chiralpak AD showed more or less enantioseparation than the covalently bonded CSP of Chiralpak IA with the same chiral selector of amylose tris(3,5‐dimethylphenylcarbamate). However, the coated Chiralcel OD showed greater enantioseparation than the covalently bonded Chiralpak IB with the same chiral selector of cellulose tris(3,5‐dimethylphenylcarbamate). Among all examined CSPs, Chiralcel OD afforded the greatest performance for enantiomer resolution of N‐PHT α‐amino acids and, therefore, all analytes enantiomers were baseline separated on Chiralcel OD. The chromatographic method developed in this study was usefully applied for determination of the enantiomeric purity of commercially available N‐PHT α‐amino acids analytes.

A weak anion-exchanger chiral selector, quinine-incorporated silica/zirconia hybrid monolithic (QUI-S/ZHM) capillary column was prepared by sol–gel technology. The performance of the QUI-S/ZHM column was investigated for enantioresolution of a set of acidic chiral drugs and dinitrobenzoyl (DNB)-amino acids by capillary electrochromatography in aqueous organic mobile phases composed of acetonitrile (ACN) and triethylammonium acetate (TEAA) buffer. Effects of several parameters including the ACN content, concentration and pH of the mobile phase on the chiral separation were examined. Baseline resolutions of all the compounds were obtained in the mobile phase consisting of 70:30 ACN/TEAA (10 mM, pH 6) under applied voltage of −10 kV at 25 °C within 20 min.

Liquid chromatographic comparisons for enantiomer resolution of N‐fluorenylmethoxycarbonyl (FMOC) α‐amino acids with fluorescence detection were made on covalently bonded type chiral stationary phases (CSPs) (Chiralpak IA and Chiralpak IB) and coated type CSPs (Chiralpak AD and Chiralcel OD) derived from polysaccharide derivatives of the same chiral selectors. This is the first study reported of enantiomer resolution with fluorescence detection on covalently bonded type CSPs, Chiralpak IA and Chiralpak IB. In general, covalently bonded type CSPs (Chiralpak IA and Chiralpak IB) showed lower enantioseparation than coated type CSPs (Chiralpak AD and Chiralcel OD) for enantiomer resolution of these analytes, respectively. Owing to higher sensitivity and broader solvent compatibility in fluorescence detection on Chiralpak IA and Chiralpak IB than in UV detection, however, this analytical method is expected to enlarge their application of enantiomer resolution, such as an online HPLC monitoring of asym...

Although chiral separation has been one of the main topics of chromatographic practice for over twenty-five years, it still presents many difficulties. In this work, the ultrasonic dependence of chiral resolution was investigated at various temperatures to improve resolution and reduce analysis time. The chiral resolution Was performed on recently commercialized two HPLC chiral stationary phases (CSP 1 and CSP 2) with the analogues of racemic N-acylnaphthylethylamines (la-d) and racemic amino acid derivatives (2a-c, 3a-c) as analytes. The CSP 1 was prepared from a (R)-N-(3,5-dinitrobenzoyl)phenylglycinol and the CSP 2 was prepared from a (S)-N-3,5-(dinitrobenzoyl) leucine. From the comparison of the chromatographic results under sonic condition with those under non-sonic condition, we found that the ultrasound decreased the elution time in chiral chromatography at all temperatures and improved the enantioselectivity at high temperature (45, 50, 60 °C).

We report use of β-cyclodextrin (CD)-hexamethylene diisocyanate (HMDI) copolymer(CDPU)-coated zirconia (CDPUZ) as a chiral stationary phase for separation of enantiomers of a set of 2,4-dinitrophenyl (DNP) amino acids in reversed-phase liquid chromatography. The effects of polymer loading on zirconia and HMDI/CD molar ratio in the preparation of CDPU on retention and selectivity in the separation of DNP-amino acids in were examined to find optimum polymer loading and HMDI/CD ratio. It was observed that 8% loading of the CDPU prepared with the HMDI/CD molar ratio of 4 gave the best separation of the amino acids investigated. Retention and chiral selectivity of CDPUZ were also compared to those for previously reported carboxymethyl-β-CD-coated zirconia (CMCDZ). CDPUZ gave better chiral separation than CMCDZ for the DNP-amino acids.

Quinine has been widely used for enantiomer separation in HPLC as a chiral ion-pairing agent in the mobile phases and as a CSP ligand. In recent years, chiral stationary phases (CSPs) based on the use of carbamoylated derivatives of quinine and quinidine as selectors were found to be highly stereoselective for the direct resolution of chiral acids using mixtures of aqueous buffers and methanol or acetonitrile as mobile phases. Silica is the most popular choice for support of HPLC stationary phase ligands due to the mechanical strength, wide range of particle size and pore dimensions, pore structure and well-established silane chemistry. However, silica and bonded phase ligands have stability problems. Silica dissolves in mobile phase buffered at or above pH 8 with loss of bonded phase ligand and column packing. Loss of organosilanes from the silica surface via hydrolysis proceeds rapidly at low pH (< 3) and at high temperature (≥ 40 C). These deficiencies of the column packing create problems of poor injection reproducibility, poor peak shape, and high backpressure, thus making method development tasks difficult. Zirconia particles are very robust material; they show no detectable signs of dissolution over the pH range from 1 to 14 and have been used for prolonged periods at temperatures up to 200 C in chromatographic separations. Over the last decade, zirconia has received considerable attention as stationary phase support for HPLC. We have been working to develop efficient and chemically stable CSPs on zirconia substrate. Bare zirconia cannot be covalently modified like silica due to the instability of Zr-C and Zr-OSi bonds in water. Zirconia-based CSPs reported have thus been prepared by coating chiral selectors on zirconia surface by utilizing Lewis acid-base chemistry. Recently we reported chiral separation of N-(2,4-dinitrophenyl) (DNP) amino acids on a 9-O-(phenyloxycarbonyl)quinine bonded carbon-clad zirconia (QNOCZ) in reversedphase liquid chromatography (RPLC). Carbon-clad zirconia is made by passing organic vapors over very hot porous zirconia. Carbon-clad zirconia particles show similar mechanical, thermal and chemical stability to bare zirconia particles but no appreciable Lewis acidity. They do not exhibit peak tailing for amines nor do they adsorb phosphates or carboxylates. Although QNOCZ gave better chiral selectivity for amino acids studied than quinine-coated zirconia and quinine-bonded silica, synthetic yield of the chiral selector, 9-O-(4-aminophenyloxycarbonyl)quinine was very low (14%) and bonding yield of the selector to carbon-clad zirconia was also found to be also very low due to several intermolecular site reactions of the diazonium salt of the selector. In this work we prepared 9-O-(phenylcarbamoyl)quininebonded carbon-clad zirconia (QNCCZ) in the hope of obtaining better synthetic and bonding yields than those for QNOCZ. It was also expected that different functionality in the linkage (carbamoyl vs. oxycarbonyl) would show different retention and selectivity behavior. QNCCZ was used as the chiral stationary phase (CSP) for separation of enantiomers of DNP-amino acids in RPLC. Retention and enantioselectivity for QNCCZ were compared to those for QNOCZ and quinine carbamate-bonded silica (QNS).

In a number of previous communications, we reported on the utility of the solvatochromic linear solvation energy relationship (LSER) method for the correlation of a number of solute and solvent-dependent properties. In those studies, it was our practice to examine the effect of a variety of solvents on a given solute or a number of solutes in a given solvent. Here we report on a novel generalized LSER in which the solute and solvent were both simultaneously varied so as to assess the validity of the entire LSER concept and define its limits. The Hildebrand solubility parameter, δ H , the Kamlet–Taft solvatochromic parameters, π *, α , β and the solute molar volume, V 2 , were used as the explanatory variables. The gas–liquid partition coefficient ( K ) was the property of interest. We have found that the correlation using the generalized linear solvation energy relationship is statistically as good as the previous LSER correlations despite the use of a far smaller number of freely adjustable parameters. Furthermore, the new approach is able to give reasonable predictions of K values of systems not included in the data set upon which the regression is based.

Zirconia is known to be one of the best chromatographic support materials due to its excellent chemical, thermal, and mechanical stability. A quinine carbamate-coated zirconia was prepared as a chiral stationary phase for separation of enantiomers of DNP-amino acids in reversed-phase liquid chromatography. Retention and enantioselectivity of this phase were compared to those for quinine carbamate bonded onto silica. Most amino acids studied were separated on the quinine carbamate-zirconia CSP although retention was longer and chiral selectivity was somewhat lower than on the corresponding silica CSP. Increased retention and decreased selectivity are probably due to strong non-enantioselective Lewis acid-base interactions between the amino acid molecule and the residual Lewis acid sites on the zirconia surface.

Zirconia particles are a very robust material and have received considerable attention as a stationary phase support for HPLC. Chromatographic performances of ADMPC- and CDMPC-coated zirconia as chiral stationary phases (CSPs) were compared by measuring the chiral discrimination of a set of 14 racemic compounds on the two columns. On the whole CDMPC-zirconia showed better chiral selectivity for the test compounds than ADMPC-zirconia. The two CSPs show complementary chiral recognition capability for the types of racemates studied. Chiral selectivities of ADMPC- and CDMPC-coated zirconia were also compared with those of ADMPC- and CDMPC-coated silica. The zirconia CSPs exhibited comparable and sometimes better enantioselectivity than the corresponding silica CSPs.

It is well-known in reversed-phase liquid chromatography that the solute retention factor (k) in binary eluents is well modeled as a quasi-linear function of the eluent composition (Φ) by the equat...

Norfloxacin, that inhibits the action of topoisomerase II, binds to wide variety of DNA. The binding mode of this drug to double- and super-coiled DNA (ds- and scDNA) is compared in this study by various spectroscopic methods, including absorption, fluorescence, and circular dichroism(CD) spectroscopy. Hypochromism in the absorption band, negative and positive induced CD bands (respectively in 240-260 nm and 270-300 nm region) are apparent for the norfloxacin that bound to both the dsDNA and scDNA. A decrease in fluorescence is also noticed in the presence of both DNAs. Since the spectroscopic characteristics are the same for both complexes, it is imperative that the binding mode of the norfloxacin is similar in ds- and scDNA. In the presence of Mg 2 + , which is a cofactor in the topoisomerase II action, the fluorescence intensity of the scDNA-norfloxacin complex increased and the resulting fluorescence intensity and shape was identical to that in the absence of scDNA. Therefore, the addition of an excess amount of Mg 2 + may result in the extrusion of norfloxacin from scDNA.

Dept. of Chemistry, University of Minnesota, MN 55455, USAReceived August 2, 2002Key Words : Chiral stationary phase, Amylose and cellulose tris(3,5-dimethylphenylcarbamate), Zirconia,HPLCHPLC separation method based on chiral stationary phases(CSPs) has become one of the most attractive approaches tochiral separations, due to their simplicity for determiningoptical purity and easy extension to the semipreparative andpreparative scales.

Poly(ethylene glycol) (PEG) in water is known to alter the structure and/or state of water to give a different polarity from that of pure water. We determined using the solvatochromic comparison method the dipolarity/polarizability (pi*), hydrogen bond (HB) accepting basicity (beta) and HB donating acidity (alpha) of aqueous solutions of PEGs of variegated molecular weights at different concentrations in order to understand the influence of the polymer on these properties of water. It was observed that PEG decreases alpha for water while it does not change pi* and beta appreciably in the range of the molecular weight and compositions studied.

An efficient synthesis of 2H-pyrans is achieved by indium (III) chloride-catalyzed reactions of 1,3-dicarbonyl compounds with a variety of α,β-unsaturated aldehydes in moderate yields. This method has been applied to the synthesis of pyranocoumarins, pyranophenalenones, and pyranoquinolinone alkaloids.

Cyclodextrin (CD) polymers were synthesized from the reaction of native CDs with a hexamethylene diisocyanate (HDI) compound in a dried DMF solution. The obtained CD polymer contained a range of 8 - 14% N due to HDI by elemental analysis. The physical and chemical properties of the CD polymers were characterized by IR, solid state 13C NMR, TGA, and DSC, respectively. An HPLC column was prepared using the CD polymer with a carbamate linker by a slurry method. Separation of the phenol isomers was conducted using the CD polymer stationary phase and CD by HPLC. Furthermore, an inclusion complex of the phenol isomer was studied by FT-Raman spectroscopy. From the HPLC and FT-Raman results, inclusion phenomena of o-, m-, and p-nitrophenol onto CD and CD polymers were analyzed.

An organic–inorganic silica/zirconia hybrid monolithic capillary column was prepared by sol–gel process in a fused-silica capillary by using triethoxysilylpropylcarbamate (TEOSPC) derivative of clindamycin phosphate (CLIP) as a chiral selector. A sol solution consisting of 6 × 10 −3 M of polyethylene glycol, 1 M of water, 2 M of acetic acid and 0.04/0.96 ratio of CLIP-TEOSPC/Zr-Bu resulted in homogeneous monolith having well defined through-pores and tightly anchored to the capillary wall. The column was employed for capillary electrochromatographic enantioseparation of eight basic chiral drugs in mobile phases consisting of acetonitrile, methanol and ammonium acetate (AA, as the electrolyte). Effects of the compositions of solvents and electrolyte in the mobile phase, applied voltage and capillary temperature on chiral separation were investigated. The highest resolution values were obtained with mobile phases consisting of 40/60 MeOH/ACN and 100 mM AA (for citalopram, Troger's base, indapamide, metoprolol, cetirizine and atropine) and 35/65 MeOH/ACN and 100 mM AA (for sertraline and propranolol) using −10 kV applied voltage at 25 °C.

We report preparation and use of carboxymethyl-β-cyclodextrin-coated zirconia as a chiral stationary phase (CSP) for separation of enantiomers of 2,4-dinitrophenyl (DNP) amino acids in reversed-phase high performance liquid chromatography. The CSP showed good enantioselectivity for some of the amino acids studied. Effects of pH and amount of methanol in mobile phases on retention and enantioselectivity for the analytes were examined.

The 3,5-dinitrobenzoyl (±)-amino acid esters were successfully resolved on (−)-phenylurea chiral stationary phases (CSPs) in a normal phase mode by high-performance liquid chromatography (HPLC). The alcohols used for esterification were methanol, ethanol, and n-propanol. The effects of esterification were studied via retention and optical resolution. The solvent and its concentration effect on enantioselectivity have been investigated based on the binary or ternary solvent system. The alcohol used in the binary or ternary solvent system was critical to the enantiomeric resolution of 3,5-dinitrobenzoyl amino acid esters while the nonalcoholic solvent was not suitable. The optical condition of the enantiomeric resolution is discussed in terms of the solvent composition and structure of the amino acid esters. The main chiral recognition mechanism based on the π–π interaction of the nitrobenzoyl group of the amino acid derivatives with the π-basic phenyl group of CSPs is described.

A chiral stationary phase (CSP 2) was prepared by connecting N-(3,5-dinitrobenzoyl)-(R)-phenylglycinol to silica gel through carbamate linkage. The CSP was applied in resolving racemic N-acyl-1-arylaminoalkanes and the chromatographic resolution results were compared with those on the previously reported CSP (CSP 1) derived from N-(3,5-dinitrobenzoyl)-(2S,3R)-norephedrine. From the comparison of the resolution results on CSP 1 and CSP 2, it was concluded that CSP 2 is more effective than CSP 1 in the resolution of racemic N-acyl-1-arylaminoalkanes.

The effect of triethylamine (TEA) in the mobile phase on the RPLC retention behavior of small organic solutes has been studied on a conventional polymeric octadecylsilica (ODS) and on a horizontally polymerized ODS. Retention factors for a set of solutes were measured on the two phases with methanol-water mobile phases containing triethylamine at different concentrations and analyzed by use of linear solvation energy relationships (LSER). Variation of the resulting LSER coefficients—v (hydrophobicity),r (polarizability),s (dipolarity),b (hydrogen-bond (HB) donating acidity), anda (HB accepting strength)—were examined to see how TEA affects the intermolecular interaction properties of the mobile and stationary phases and hence the retention of the solutes. Addition of TEA to the mobile phase changes the interaction properties of both conventionally polymerized and horizontally polymerized ODS; the effect is greater for the conventional phase. The HB donating acidity (b) of conventional polymeric ODS is significantly reduced by addition of TEA. For the mobile phases studied the magnitudes of theb andv coefficients for the horizontally polymerized ODS phase are greater than for the conventional phase. The different interaction properties of the two polymeric phases arise mainly as a result of differential adsorption of TEA, because of the very different amounts of surface silanol groups present on the two phases.

Reversed-phase liquid chromatography (RPLC) separation factors for racemic 2,4-dinitrophenyl (DNP) amino acids (Phe and Trp) having various substituents at different positions were measured on native β-CD and heptakis(3-O-methyl)-β-CD as the chiral stationary phase in order to compare enantioselectivity of these phases. Both native β-CD and heptakis(3-O-methyl)-β-CD showed good enantioselectivity for the DNP–amino acids investigated. Enantioselectivities of the two CD phases for the DNP–amino acids vary with the type and position of the substituent on the DNP moiety of the amino acids. Heptakis(3-O-methyl)-β-CD, the cavity of which is more electron-rich than that of native β-CD, showed in general much better enantioselectivity for the amino acid derivatives studied.

Characterization of a number of normal-phase liquid chromatography (NP-LC) stationary phases was attempted by estimating characteristic interaction constants for the stationary phases based on linear solvation energy relationships. Five characteristic interaction constants of the stationary phases, the lipophilicity (v), polarizability (r), dipolarity (s), hydrogen bond (HB) acceptor basicity (a) and HB donor acidity strength (b) were determined by multiple regression analyses of logarithmic capacity factors (k′) for a set of test solutes measured on them in the mobile phase of a given composition vs. the solute properties represented by characteristic molecular volume (Vx), excess polarization (R2), dipolarity/polarizability (π*), HB donor acidity (α) and HB acceptor basicity (β). Magnitudes of the five constants vary with the type of the stationary phases. The effect of polar modifier in the mobile phases on the interaction properties of the sorbents was also investigated.

Recently, the solvatochromic comparison method was successfully applied to determine the effects of adding a modifier to the mobile phase in packed capillary columns, and we have extended this method to understand the modifier effect in open tubular capillary column supercritical fluid chromatography (SFC). SFC retention data for a wide variety of compounds at different modifier concentrations were obtained using a 10 m×50 μm I.D. SB-methyl-100 polydimethylsiloxane capillary column. Supercritical carbon dioxide at a pressure of 260 atm was used as mobile phase and methanol as modifier. The LSER regression equation used was log k ′=SP 0 + l log L 16 + s π 2 H + a α 2 H + b β 2 H + r R 2 , where k ′ represents the capacity factor and L 16 represents the partition coefficient of the solute between the gas phase and n -hexadecane at 298 K, and l , s , a , b and r are the regression coefficients. For open tubular column SFC, variation of the l and s coefficients showed a similar pattern to the packed capillary column SFC. However, the b coefficient showed a different pattern from packed capillary SFC, i.e., a gradual decrease with increasing modifier concentration. Based on these observations, the effects of a small amount of a modifier in the supercritical mobile phase and its effects on solute retention mechanisms have been discussed. For open tubular column SFC, the swelling of the chromatographic stationary phase by a modifier and carbon dioxide and the increase of dipolarity of the mobile phase are the most important contributing factors in substantiating the modifier effect on solute retention.

Three new β-cyclodextrin derivatives, heptakis(6-O-isopropyldi-methylsilyl-2,3-di-O-ethyl)-β-cyclodextrin, heptakis(6-O-thexyldi-methylsilyl-2,3-di-O-ethyl)-β-cyclodextrin, and heptakis(6-O-cy-clohexyldimethyl-2,3-di-O-ethyl)-β-cyclodextrin (IPDE-β-CD, TXDE-β-CD, and CHDE-β-CD), were synthesized and the enan-tioselectivities of these three CD derivatives and heptakis(6-O-tert-butyldimethylsilyl-2,3-di-O-ethyl)-β-cyclodextrin (TBDE-β-CD) were compared for GC separation of a range of chiral test com-pounds. In particular TXDE-β-CD showed much higher enentio-selectivity than TBDE-β-CD. Enentioselectivities of IPDE-β-CD and CHDE-β-CD are somewhat lower than that of TXDE-β-CD and CHDE-β-Cd are somewhat lower than that of TXDE-β-CD. These observations are indicative of significant effects of subtle changes in the structure of the 6-O-substituent on the enantioselec-tivity of the β-CD derivatives. The difference in enantioselectivities of the 6-O-substituted CD derivatives were explained in terms of relative contributions of the effects of hydrophobicity and steric hindrance of the substituent to the inclusion process. CHDE-β-CD showed the lowest enantioselectivity among the threederivatives. It is likely that the unfavorable steric hindrance of the bulky cyclo-hexyl group plays a greater role than the favorable hydrophobicity effect of the cyclohexyl group in the inclusion process in CHDE-β-CD. IPDE-β-CD showed lower selectivity than TXDE-β-CD and TBDE-β-CD. In the case of these CD derivatives having acyclic substituents the relative hydrophobicity of the substituent seems to be a dominant factor affecting the inclusion process. Isopropyl groups factor affecting the inclusion process. Isopropyl groups are less hydrophobic than thexyl and tert-butyl groups.

Three new chiral selectors, 6-tert-butyldimethylsilyl-2,3-diethyl-a-cyclodextrin, 6-tert-butyldimethylsilyl-2,3-diethyl- and dipropyl-β-cyclodextrin (TBDE-α-CD, TBDE-β-CD, TBDP-β-CD) were synthesized and tested as chiral stationary phases in capillary gas chromatography. TBDE-β-CD in particular showed a high enan-tioselectivity for test chiral compounds due to good solubility in a polar polysiloxane (OV-1701). Enantioselectivity obtained with TBDE-β-CD was compared with that of 6-tert-butyldimethylsilyl-2,3-di-O-methyl-β-cyclodextrin (TBDM-β-CD). Better enantiose-lectivity was obtained with TBDE-P-CD than with TBDM-β-CD for the test chiral compounds studied. This is probably due to greater effect of the increased hydrophobicity of TBDE-β-CD which favors inclusion of the analytes than the effect of increased steric hindrance. With TBDP-β-CD the less polar lactones are well separated due most likely to increased hydrophobicity of the propyl groups while the more polar are not well resolved. For TBDP-β-CD it is likely that the unfavorable steric hindrance is predominant over the favorable hydrophobicity of the propyl groups, thus hindering the formation of inclusion complexes of the alcohols with TBDP-β-CD. TBDE-α-CD was also a valuable chiral selector for the separation of small chiral molecules such as simple secondary alcohols and nitro-substituted alcohols.

The partition and adsorption mechanisms of retention in reversed-phase liquid chromatography have been examined based on a comparison of the free energy of transfer of methylene groups from aqueous-organic mixtures to bulk hexadecane with those to a variety of polymeric bonded phases. The stationary phases studied include: conventional silica-based polymeric phases of various alkyl chain lengths, a so-called “horizontally polymerized” octadecyl phase on silica and a series of polybutadiene-coated zirconia phases. The data indicate that for methylene groups a partition-like mechanism is dominant on all phases. On the polybutadiene-coated zirconia and “horizontally polymerized” octadecyl phases the partition mechanism holds at all mobile phase compositions. In contrast on conventional polymeric silica phases the retention mechanism seems to become more adsorption-like at methanol compositions greater than about 70% (v/v).