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Retention Factors (retention + factor)
Selected AbstractsCompositional effects on electrophoretic and chromatographic figures of merit in electrokinetic chromatography with cetyltrimethylammonium bromide/sodium octyl sulfate vesicles as the pseudostationary phase.ELECTROPHORESIS, Issue 5 2008Part 1: Effect of the phase ratio Abstract The effect of the phase ratio on the electrophoretic and chromatographic properties of unilamellar vesicles comprised of cetyltrimethylammonium bromide (CTAB) and sodium octyl sulfate (SOS) was investigated in EKC. The surfactant concentration of the vesicles was 0.9, 1.2, 1.5, and 1.8% w/v, with a mole ratio of 1:3.66 (CTAB/SOS). Results were compared to those obtained using SDS micelles at concentrations of 1.0% (w/v, 35,mM) and 1.5% (52,mM). The CTAB/SOS vesicles (0.9,1.8% w/v) provided a significantly larger elution range (5.7,,,tves/t0,,,8.7) and greater hydrophobic (methylene) selectivity (2.8,,,,CH2,,,3.1) than SDS micelles (3.1,,,tmc/t0,,,3.3; ,CH2,=,2.2). Whereas the larger elution range can be attributed to the 25% reduction in EOF due to the interaction of unaggregated CTAB cations and the negatively charged capillary wall, the higher methylene selectivity is likely due to the lower concentration of water expected in the CTAB/SOS vesicle bilayer compared to the Palisades layer of SDS micelles. For a given phase ratio, CTAB/SOS vesicles are somewhat less retentive than SDS micelles, although retention factors comparable to those observed in 1.0,1.5% SDS can be obtained with 1.5,1.8% CTAB/SOS. A linear relationship was observed between phase ratio and retention factor, confirming the validity of the phase ratio model for these vesicles. Unique polar group selectivities and positional isomer shape selectivities were obtained with CTAB/SOS vesicles, with both types of selectivities being nearly independent of the phase ratio. For four sets of positional isomers, the elution order was always para < ortho < meta. Finally, the thermodynamics of solute retention was qualitatively similar to that reported for other surfactant aggregates (micelles and microemulsions); the enthalpic contribution to retention was consistently favorable for all compounds, whereas the entropic contribution was favorable only to hydrophobic solutes. [source] Low-temperature bath/coupled-capillary/sweeping-micellar electrokinetic capillary chromatography for the separation of naphthalene-2,3-dicarboxaldehyde-derivatized dopamine and norepinephrineELECTROPHORESIS, Issue 11 2005Chun-Min Shih Abstract The use of a low-temperature (0°C) bath-assisted coupled capillary for the separation of naphthalene-2,3-dicarboxaldehyde (NDA)-derivatized dopamine and norepinephrine using the sweeping-micellar electrokinetic capillary chromatography (MEKC) mode is described. In this technique, a capillary consisting of two portions with different inside diameters is used. Therefore, the field strength inside the capillary is different. Hence, the electrophoretic migration velocities of the analytes and the electroosmotic flow (EOF) also are different. Furthermore, when a portion of the capillary (wide portion, used for sweeping) is immersed in a low-temperature bath, the viscosity of the buffer and the retention factor of the analytes inside are increased. Thus, not only are the interactions between the SDS micelles and the analytes increased, but the SDS-analytes also move more slowly. As a result, a more complete separation can be achieved, even when the sample injection volume is large, up to , 2 ,L. In general, when the volume of an injected sample is larger, the effects of sweeping and separation would become insufficient, especially when the retention values (k) of the analytes are quite different. However, this limitation can be improved when the low-temperature bath/coupled capillary/sweeping-MEKC mode is used. [source] Microemulsion electrokinetic chromatography of drugs varying in charge and hydrophobicity: I. Impact of parameters on separation performance evaluated by multiple linear regression modelsELECTROPHORESIS, Issue 1 2004Valérie Harang Abstract The separation of anionic, cationic and neutral drugs in microemulsion electrokinetic chromatography (MEEKC) was studied with a statistical experimental design. The concentration of sodium dodecyl sulfate (SDS, surfactant), 1-butanol (co-surfactant) and borate buffer and the factors Brij 35 (surfactant), 2-propanol (organic solvent) and cassette temperature were varied simultaneously, while the parameters pH (9.2), the concentration of octane (oil, 0.8% w/w), the voltage (10 kV) and the dimension of the fused-silica capillary, were kept constant. Eight different model substances were chosen with different hydrophobicities. Two of the analytes were positively charged, two were negatively charged, and the remaining four were neutral or close to neutral at the pH explored. The importance of each parameter on the separation window, the plate height and the retention factor for each of the analytes was studied by means of multiple linear regression (MLR) models. A new response was evaluated for anions, the quotient between the effective mobility in the microemulsion and the effective mobility in the corresponding buffer. Factors affecting selectivity changes were also explored, and it was found that SDS and 2-propanol had the largest effect on selectivity. [source] Paralog of the formylglycine-generating enzyme , retention in the endoplasmic reticulum by canonical and noncanonical signalsFEBS JOURNAL, Issue 6 2008Santosh Lakshmi Gande Formylglycine-generating enzyme (FGE) catalyzes in newly synthesized sulfatases the oxidation of a specific cysteine residue to formylglycine, which is the catalytic residue required for sulfate ester hydrolysis. This post-translational modification occurs in the endoplasmic reticulum (ER), and is an essential step in the biogenesis of this enzyme family. A paralog of FGE (pFGE) also localizes to the ER. It shares many properties with FGE, but lacks formylglycine-generating activity. There is evidence that FGE and pFGE act in concert, possibly by forming complexes with sulfatases and one another. Here we show that human pFGE, but not FGE, is retained in the ER through its C - terminal tetrapeptide PGEL, a noncanonical variant of the classic KDEL ER-retention signal. Surprisingly, PGEL, although having two nonconsensus residues (PG), confers efficient ER retention when fused to a secretory protein. Inducible coexpression of pFGE at different levels in FGE-expressing cells did not significantly influence the kinetics of FGE secretion, suggesting that pFGE is not a retention factor for FGE in vivo. PGEL is accessible at the surface of the pFGE structure. It is found in 21 mammalian species with available pFGE sequences. Other species carry either canonical signals (eight mammals and 26 nonmammals) or different noncanonical variants (six mammals and six nonmammals). Among the latter, SGEL was tested and found to also confer ER retention. Although evolutionarily conserved for mammalian pFGE, the PGEL signal is found only in one further human protein entering the ER. Its consequences for KDEL receptor-mediated ER retrieval and benefit for pFGE functionality remain to be fully resolved. [source] Retention of arsenic species on zwitterionic stationary phase in hydrophilic interaction chromatographyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 6-7 2010Dan Xie Abstract Zwitterionic hydrophilic interaction chromatography (ZIC® -HILIC) was used to study the retention of selected organoarsenicals. The retention behavior of nine organic arsenic species on ZIC® -HILIC was investigated to elucidate which is the driving force for their separation, hydrophilic partitioning or adsorption driven by hydrogen bonds with surface H-donor/acceptor groups of the stationary phase. For this, the retention factor of the compounds k was correlated with log PO/W and with the calculated strength of hydrogen bonding of the analytes. By examining aliphatic and phenylic compounds separately, improved correlation was received. This indicates that both phenomena contribute to the separation of these arsenic species on ZIC® -HILIC. The results obtained evidence that considerable electrostatic interactions also occur on ZIC® -HILIC. Retention behavior of arsenic species was investigated by varying the separation conditions, which shows that the composition of the eluent has a strong influence on the retention behavior. It is highly dependent on water/acetonitrile ratio, pH value and salt additives. Dissociation degree and polarity of arsenic species, which are varying with pH, regulate the distribution of arsenic species between stationary and mobile phases in HILIC. Increase in the ammonium acetate concentration leads to shortened or to prolonged retention depending on the structure of the arsenic species. [source] Visible light initiated polymerization of styrenic monolithic stationary phases using 470,nm light emitting diode arraysJOURNAL OF SEPARATION SCIENCE, JSS, Issue 1 2010Zarah Walsh Abstract Poly(styrene- co -divinylbenzene) monolithic stationary phases have been synthesized for the first time by photoinitiated polymerization. An initiator composed of (+)-(S)-camphorquinone/ethyl-4-dimethylaminobenzoate/N- methoxy-4-phenylpyridinium tetrafluoroborate was activated using a 470,nm light emitting diode array as the light source. Spatially controlled polymerization of styrenic monoliths has been achieved within specific sections of a 100,,m id polytetrafluoroethylene-coated fused-silica capillary using simple photo masking. The sharpness of the edges was confirmed by optical microscopy, while SEM was used to verify a typical porous, globular morphology. Flow resistance data were used to assess the permeability of the monoliths and they were found to have good flow through properties with a flow resistance of 0.725,MPa/cm at 1,,L/min (water, 20°C). Conductivity profiling along the length of the capillary was used to assess their lateral homogeneity. Monoliths which were axially rotated during polymerization were found to be homogeneous along the whole length of the capillary. The monolithic stationary phases were applied to the RP gradient separation of a mixture of proteins. Column fabrication showed excellent reproducibility with the retention factor (k) having a RSD value of 2.6% for the batch and less than 1.73% on individual columns. [source] A computational study of the porosity effects in silica monolithic columnsJOURNAL OF SEPARATION SCIENCE, JSS, Issue 10-11 2004Piotr Gzil Abstract We report on a theoretical study of the influence of the through-pore porosity on the main chromatographic performance parameters (reduced theoretical plate height, flow resistance, and separation impedance) of silica monoliths. To investigate this problem devoid of any structural uncertainties, computer-generated structural mimics of the pore geometry of silica monolithic columns have been studied. The band broadening in these synthetic monoliths was determined using a commercial Computational Fluid Dynamics (CFD) software package. Three widely differing external porosities (, = 0.38, , = 0.60, and , = 0.86) are considered and are compared on the basis of an identical intra-skeleton diffusivity (D s = 5×10,10m2/s), internal porosity (,int = 0.5), and for the same phase retention factor (k ´ = 1.25). Since the data are obtained for perfectly ordered structures, the calculated plate heights and separation impedances constitute the ultimate performance ever to be expected from a monolithic column. It is found that, if silica monoliths could be made perfectly homogeneous, domain size-based reduced plate heights as small as h min , 0.8 (roughly independent of the porosity) and separation impedances as small as Emin , 130 (, = 0.60) and Emin , 40 (, = 0.86) should be achievable with pure water as the working fluid. The data also show that, although the domain size is a much better reduction basis than the skeleton size, the former is still not capable of bringing the van Deemter curves of different porosity columns into perfect agreement in the C term dominated velocity range. It is found that, in this range, large porosity monoliths can be expected to yield smaller domain size-based reduced plate heights than small porosity monoliths. [source] Chromatographic classification and comparison of commercially available perfluorinated stationary phases for reversed-phase liquid chromatography using Principal Component AnalysisJOURNAL OF SEPARATION SCIENCE, JSS, Issue 3-4 2003Melvin R. Euerby Abstract A range of ten perfluorophenyl and perfluoroalkyl stationary phases has been evaluated using standard chromatographic tests and probes. Principal Component Analysis of the data has indicated that the phases can be divided into distinct groupings. Extending the dataset to include standard alkyl and phenyl phases provided further data interpretation to support the orthogonal selectivity claims made for perfluorinated phases. The analysis of a range of basic analytes showed an unusual extended retention of hydrophilic basic analytes with perfluorophases. Furthermore, a non-linear relationship between the amount of organic modifier and the logarithm of the retention factor was observed, for the hydrophilic bases, which could not be modelled with LC prediction softwares. This was in sharp contrast to the alkyl and phenyl phases examined. Basic analyte retention on perfluoroalkyl phases could be modelled adequately for the lipophilic bases. Exploration of the retention mechanism of these perfluoro phases indicated that silanol interactions were important in retention and selectivity. Using a rapid, isocratic, high organic modifier methodology, it was possible to analyse a mixture containing a lipophilic steroid, hydrophilic base and an internal standard in < 4 minutes with a perfluorophenyl phase. This had previously only been achievable with an alkyl phase under gradient elution conditions. [source] Phospholipids in liquid chromatography/mass spectrometry bioanalysis: comparison of three tandem mass spectrometric techniques for monitoring plasma phospholipids, the effect of mobile phase composition on phospholipids elution and the association of phospholipids with matrix effectsRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 14 2009Yuan-Qing Xia Because plasma phospholipids may cause matrix effects in bioanalytical liquid chromatography/tandem mass spectrometry (LC/MS/MS) methods, it is important to establish optimal mass spectrometric techniques to monitor the fate of phospholipids during method development and application. We evaluated three MS/MS techniques to monitor phospholipids using positive and negative electrospray ionization (ESI). The first technique is based on using positive precursor ion scan of m/z 184, positive neutral loss scan of 141 Da and negative precursor ion scan of m/z 153. The second technique is based on using class-specific positive and negative selected reaction monitoring (SRM) transitions to monitor class-representative phospholipids. The third technique, previously reported, utilizes in-source collision-induced dissociation (CID)-based positive SRM of m/z 184,,,184. We recommend the all-inclusive technique 1 for use in qualitative assessment of all classes of phospholipids and technique 2 for use in quantitative assessment of class-representative phospholipids. Secondly, we evaluated the elution behaviors of the plasma phospholipids under different reversed-phase mobile phase conditions. The phospholipid-eluting strength of a mobile phase was mainly dependent on the type and amount (%) of the organic eluent and the strength increased in the order of methanol, acetonitrile and isopropyl alcohol. Under the commonly used gradient and isocratic elution schemes in LC/MS/MS bioanalysis, not all the phospholipids are eluted off the column. Thirdly, we investigated the association between phospholipids and matrix effects in positive and negative ESI using basic, acidic and neutral analytes. While the phospholipids caused matrix effects in both positive and negative ESI, the extent of ionization suppression was analyte-dependent and was inversely related to the retention factor and broadness of the phospholipids peaks. The lysophospholipids which normally elute earlier in reversed-phase chromatography are more likely to cause matrix effects compared to the later-eluting phospholipids in spite of the larger concentrations of the latter in plasma. Copyright © 2009 John Wiley & Sons, Ltd. [source] Study of influence of additives of tyloxapol on the chromatographic characteristics of the model compounds: the comparative characterization of micellar mobile phases of tyloxapol and Triton X-100BIOMEDICAL CHROMATOGRAPHY, Issue 7 2009Marina Rukhadze Abstract The chromatographic behavior of model compounds of biomedical significance (organic acids, amino acids, drugs) was investigated using mobile phases modified with tyloxapol. The influence of factors such as concentration of tyloxapol, content of organic modifier and pH of mobile phase on the retention factor of solutes was studied. The results were compared with the data obtained by elution with mobile phases containing Triton X-100 additives, since units of Triton X-100 are repeated in the structure of tyloxapol. Divergence in chromatographic behavior of model compounds was explained by the difference in physico-chemical characteristics (microviscosity, polarity, critical micelle concentration, shape of micelles, etc.) of tyloxapol and Triton X-100 micelles. Copyright © 2009 John Wiley & Sons, Ltd. [source] True and Apparent Temperature Dependence of Protein Adsorption Equilibrium in Reversed-Phase HPLCBIOTECHNOLOGY PROGRESS, Issue 6 2002Szabelski The adsorption behavior of bovine insulin on a C8 -bonded silica stationary phase was investigated at different column pressures and temperatures in isocratic reversed-phase HPLC. Changes in the molar volume of insulin (, Vm) upon adsorption were derived from the pressure dependence of the isothermal retention factor ( k,). The values of , Vm were found to be practically independent of the temperature between 25 and 50 °C at ,96 mL/mol and to increase with increasing temperature, up to ,108 mL/mol reached at 50 °C. This trend was confirmed by two separate series of measurements of the thermal dependence of ln( k,). In the first series the average column pressure was kept constant. The second series involved measurements of ln( k,) under constant mobile-phase flow rate, the average column pressure varying with the temperature. In both cases, a parabolic shape relationship was observed between ln( k,) and the temperature, but the values obtained for ln k, were higher in the first than in the second case. The relative difference in ln( k,), caused by the change in pressure drop induced by the temperature, is equivalent to a systematic error in the estimate of the Gibbs free energy of 12%. Thus, a substantial error is made in the estimates of the enthalpy and entropy of adsorption when neglecting the pressure effects associated with the change in the molar volume of insulin. This work proves that the average column pressure must be kept constant during thermodynamic measurements of protein adsorption constants, especially in RPLC and HIC. Our results show also that there is a critical temperature, Tc , 53 °C, at which ln( k,) is maximum and the insulin adsorption process changes from an exothermic to an endothermic one. This temperature determines also the transition point in the molecular mechanism of insulin adsorption that involves successive unfolding of the protein chain. [source] Multi-walled carbon nanotube composites with polyacrylate prepared for open-tubular capillary electrochromatographyELECTROPHORESIS, Issue 19 2010Jian-Lian Chen Abstract A new phase containing immobilized carbon nanotubes (CNTs) was synthesized by in situ polymerization of acid-treated multi-walled CNTs using butylmethacrylate (BMA) as the monomer and ethylene dimethacrylate as the crosslinker on a silanized capillary, forming a porous-layered open-tubular column for CEC. Incorporation of CNT nanomaterials into a polymer matrix could increase the phase ratio and take advantage of the easy preparation of an OT-CEC column. The completed BMA-CNT column was characterized by SEM, ATR-IR, and EOF measurements, varying the pH and the added volume organic modifier. In the multi-walled CNTs structure, carboxylate groups were the major ionizable ligands on the phase surface exerting the EOF having electroosmotic mobility, 4.0×104,cm2,V,1,S,1, in the phosphate buffer at pH 2.8 and RSD values (n=5), 3.2, 4.1, and 4.3%, for three replicate capillaries at pH 7.6. Application of the BMA-CNT column in CEC separations of various samples, including nucleobases, nucleosides, flavonoids, and phenolic acids, proved satisfactory upon optimization of the running buffers. Their optima were found in the borate buffers at pH 9.0/50,mM, pH 9.5/10,mM/50% v/v ACN, and pH 9.5/30,mM/10% v/v methanol, respectively. The separations could also be used to assess the relative contributions of electrophoresis and chromatography to the CEC mechanism by calculating the corresponding velocity and retention factors. Discussions about interactions between the probe solutes and the bonded phase included the ,,, interactions, electrostatic repulsion, and hydrogen bonding. Furthermore, a reversed-phase mode was discovered to be involved in the chromatographic retention. [source] Liposomes for entrapping local anesthetics: A liposome electrokinetic chromatographic studyELECTROPHORESIS, Issue 9 2010Jana Lokajová Abstract Bupivacaine is a lipophilic, long-acting, amide class local anesthetic commonly used in clinical practice to provide local anesthesia during surgical procedures. Several cases of accidental overdose with cardiac arrest and death have been reported since bupivacaine was introduced to human use. Recent case reports have suggested that Intralipid (Fresenius Kabi) is an effective therapy for cardiac toxicity from high systemic concentrations of, e.g. bupivacaine, even though the mechanism behind the interaction is not fully clear yet. Our long-term aim is to develop a sensitive, efficient, and non-harmful lipid-based formulation to specifically trap harmful substances in vivo. In this study, the in vitro interaction of local anesthetics (bupivacaine, prilocaine, and lidocaine) with Intralipid or lipid vesicles containing phosphatidylglycerol, phosphatidylcholine, cardiolipin, cholesterol, and N -palmitoyl- D - erythro -sphingosine (ceramide) was determined by liposome electrokinetic chromatography. The interactions were evaluated by calculating the retention factors and distribution constants. Atomic force microscopy measurements were carried out to confirm that the interaction mechanism was solely due to interactions between the analytes and the moving pseudostationary phase and not by interactions with a stationary lipid phase adsorbed to the fused-silica wall. The heterogeneity of the liposomes was also studied by atomic force microscopy. The liposome electrokinetic chromatography results demonstrate that there is higher interaction between the drugs and negatively charged liposome dispersion than with the commercial Intralipid dispersion. [source] Carboxylic multi-walled carbon nanotubes as immobilized stationary phase in capillary electrochromatography,ELECTROPHORESIS, Issue 18 2008Lorena Sombra Abstract Carboxylic multi-walled carbon nanotubes (c-MWNT) have been immobilized into a fused-silica capillary for capillary electrochromatography. The c-MWNT were successfully incorporated after the silanization and coupling with glutaraldehyde on the inner surface of the capillary. The electrochromatographic features of the c-MWNT immobilized stationary phase have been evaluated for the analysis of different compounds of pharmaceutical interest. The results indicated high electrochromatographic resolution, good capillary efficiency and retention factors. In addition, highly reproducible results between runs, days and capillaries were obtained. [source] Monomeric and polymeric anionic gemini surfactants and mixed surfactant systems in micellar electrokinetic chromatography.ELECTROPHORESIS, Issue 2 2005Part II: Characterization of chemical selectivity using two linear solvation energy relationship models Abstract Sodium di(undecenyl) tartarate monomer (SDUT), a vesicle-forming amphiphilic compound possessing two hydrophilic carboxylate headgroups and two hydrophobic undecenyl chains, was prepared and polymerized to form a polymeric vesicle (i.e., poly-SDUT). The anionic surfactants of SDUT and poly-SDUT (carboxylate head group) and sodium dodecyl sulfate, SDS (sulfate head groups) as well as mixed surfactant systems (SDS/SDUT, SDS/poly-SDUT, and SDUT/poly-SDUT) were applied as pseudostationary phases in micellar electrokinetic chromatography (MEKC). Two linear solvation energy relationship (LSER) models, i.e., solvatochromic and solvation parameter models, were successfully applied to investigate the effect of the type and composition of pseudostationary phases on the retention mechanism and selectivity in MEKC. The solvatochromic and solvation parameter models were used to help understand the fundamental nature of the solute-pseudostationary phase interactions and to characterize the properties of the pseudostationary phases (e.g., solute size and hydrogen bond-accepting ability for all pseudostationary phases). The solute types were found to have a significant effect on the LSER system coefficients and on the predicted retention factors. Although both LSER models provide the same information, the solvation parameter model is found to provide much better results both statistically and chemically than the solvatochromic model. [source] Permeation of bioactive constituents from Arnica montana preparations through human skin in-vitroJOURNAL OF PHARMACY AND PHARMACOLOGY: AN INTERNATI ONAL JOURNAL OF PHARMACEUTICAL SCIENCE, Issue 9 2006I. A. Tekko This study investigated and characterised transdermal permeation of bioactive agents from a topically applied Arnica montana tincture. Permeation experiments conducted over 48 h used polydimethylsiloxane (silastic) and human epidermal membranes mounted in Franz-type diffusion cells with a methanol-water (50:50 v/v) receptor fluid. A commercially available tincture of A. montana L. derived from dried Spanish flower heads was a donor solution. Further donor solutions prepared from this stock tincture concentrated the tincture constituents 1, 2 and 10 fold and its sesquiterpene lactones 10 fold. Permeants were assayed using a high-performance liquid chromatography method. Five components permeated through silastic membranes providing peaks with relative retention factors to an internal standard (santonin) of 0.28, 1.18, 1.45, 1.98 and 2.76, respectively. No permeant was detected within 12 h of applying the Arnica tincture onto human epidermal membranes. However, after 12 h, the first two of these components were detected. These were shown by Zimmermann reagent reaction to be sesquiterpene lactones and liquid chromatography/diode array detection/mass spectrometry indicated that these two permeants were 11,13-dihydrohelenalin (DH) analogues (methacrylate and tiglate esters). The same two components were also detected within 3 h of topical application of the 10-fold concentrated tincture and the concentrated sesquiterpene lactone extract. [source] Macroporous molecularly imprinted monolithic polymer columns for protein recognition by liquid chromatographyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 17-18 2010Jinxiang Liu Abstract Macroporous cytochrome c (cyc)-imprinted monolithic polyarylamide columns were prepared, and applied for the template protein recognition by HPLC. With cyc (18.8,mg) as template, the imprinted monolithic materials were in situ polymerized in an HPLC column tube, with methacrylamide (450,mg), methacrylic acid (15.8,mg), piperazine diacrylamide (720,mg) and ammonium sulfate (390,mg) dissolved in 5,mL of phosphate buffer (pH 7.4), initiated by ammonium persulfate and TEMED. After the reaction, cyc was removed with acetic acid (10%, v/v) containing 10%,w/v SDS. The non-imprinted monolithic column was prepared under the same procedure except without cyc. Retention of cyc and its competitive protein, lysozyme (lys), on molecular-imprinting polymer (MIP) and non-imprinted polymer columns was studied. When the pH value of mobile phase was 4.0, on MIP column, the retention factors of cyc and lys were 2.0 and 1.3, respectively. However, those on non-imprinted polymer column were very similar, both as 1.1. Even in competitive environment, a mixture of cyc and lys could be separated on MIP column under gradient elution, with resolution as 1.2. These results indicate that protein-imprinted monolithic polymer columns could offer obvious affinity and specific recognition to the template protein. [source] Preparation and characterization of polymethacrylate monolithic capillary columns with dual hydrophilic interaction reversed-phase retention mechanism for polar compoundsJOURNAL OF SEPARATION SCIENCE, JSS, Issue 15-16 2009í Urban Abstract Monolithic columns for capillary hydrophilic interaction liquid chromatography (HILIC) were prepared in fused-silica capillaries by radical co-polymerization of [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfopropyl)ammonium hydroxide and ethylene dimethacrylate in various binary and ternary porogen solvent mixtures with azobisisobutyronitrile as the initiator of the polymerization reaction. Columns showed mixed separation modes: reversed-phase (RP) in water-rich mobile phases and HILIC at high concentrations of acetonitrile (>60,80%) in aqueous,organic mobile phases. A continuous change in retention was observed at increasing concentration of water in acetonitrile, giving rise to characteristic U-turn plots of retention factors versus the concentration of water in the mobile phase, with minima corresponding to the transition between the mechanisms controlling the retention. The selectivity of organic polymer monolithic columns for HILIC separations can be varied by adjusting the concentration of sulfobetaine monomer and the composition of the porogen solvent in the polymerization mixture. Under HILIC conditions, the monolithic capillary sulfobetaine columns show separation selectivities for polar phenolic acids similar to those of a commercial silica-based sulfobetaine ZIC-HILIC column, which, however, has limited selectivity in the RP mode due to lower retention. [source] Effect of temperature on the chromatographic retention of ionizable compounds.JOURNAL OF SEPARATION SCIENCE, JSS, Issue 6-7 2008Abstract We propose a general simple equation for accurately predicting the retention factors of ionizable compounds upon simultaneous changes in mobile phase pH and column temperature at a given hydroorganic solvent composition. Only four independent experiments provide the input data: retention factors measured in two pH buffered mobile phases at extreme acidic and basic pH values (e. g., at least ± 2 pH units far from the analyte pKa) and at two column temperatures. The equations, derived from the basic thermodynamics of the acid,base equilibria, additionally require the knowledge of the solute pKa and enthalpies of acid,base dissociation of both the solute and the buffer components in the hydroorganic solvent mixture. The performance of the predictive model is corroborated with the comparison between theoretical and experimental retention factors of several weak acids and bases of important pharmacological activity, in mobile phases containing different buffer solutions prepared in 25% w/w ACN in water and at several temperatures. [source] Evaluation of a hydrazide-linked ,1 -acid glycoprotein chiral stationary phase: Separation of R - and S -propranololJOURNAL OF SEPARATION SCIENCE, JSS, Issue 10 2006Hai Xuan Abstract The binding and chiral separation of R - and S -propranolol was investigated on a new type of ,1 -acid glycoprotein (AGP) column. This column was prepared through the controlled and mild oxidation of AGP, followed by the immobilization of this protein to hydrazide-activated silica. The effects of temperature, pH, ionic strength, and organic modifiers on the retention and separation of R - and S -propranolol were investigated on this column. Both the association equilibrium constants and number of binding sites for R/S -propranolol on the AGP column were found to increase with temperature and affect the measured retention factors for these compounds. Regarding the other factors, a change in the organic modifier concentration was found to give the largest change in retention and separation. It was found through these studies that both coulombic and hydrophobic interactions played important roles in determining the retention of R - and S -propranolol on the AGP column. The efficiency and separation impedance of this system were also considered. Under the final optimum conditions identified in this study, it was possible to separate R - and S -propranolol with a resolution of greater than 1.38 in less than 5 min on a 4.1 mm I.D.×5 cm column. [source] Therapeutic monitoring of imipramine and desipramine by micellar liquid chromatography with direct injection and electrochemical detectionBIOMEDICAL CHROMATOGRAPHY, Issue 5 2005Devasish Bose Abstract A micellar liquid chromatographic (MLC) procedure was developed for the clinical monitoring of imipramine and its active metabolite, desipramine. The determination of these highly hydrophobic substances was carried out after direct injection of the serum samples using a mobile phase composed of 0.15 m SDS,6% (v/v) pentanol buffered at pH 7, pumped at 1.5 mL/min into a C18 column (250 × 4.6 mm), and electrochemical detection at 650 mV. Using this MLC method, calibration was linear (r > 0.995) and the limits of detection (ng/mL) were 0.34 and 0.24 for imipramine and desipramine, respectively. Repeatabilities and intermediate precision were tested at three different concentrations in the calibration range and a CV (%) below 2.2 was obtained. In this MLC procedure, the serum is determined without treatment, thus allowing repeated serial injections without changes in retention factors, and reducing the time and consumables required to carry out the pretreatment process. The assay method can be applied to the routine determination of serum imipramine and its metabolite in therapeutic drug monitoring. Copyright © 2004 John Wiley & Sons, Ltd. [source] Prediction of Band Profiles of Mixtures of Bradykinin and Kallidin from Data Acquired by Competitive Frontal AnalysisBIOTECHNOLOGY PROGRESS, Issue 3 2003Dongmei Zhou The competitive adsorption isotherms of two closely related peptides, bradykinin and kallidin, were measured by frontal analysis on a Zorbax SB-C18 microbore column. An aqueous soluton at 20% acetonitrile (0.1% TFA) was used as the mobile phase. The competitive isotherm data were fitted to four different models: Langmuir, Bilangmuir, Langmuir-Freundlich, and Toth. These data fitted best to a Bilangmuir isotherm model. The influence of the pressure on the retention factors of the two peptides was found to be small and was not investigated in detail. The band profiles of large samples of the single components and of their mixtures were recorded. The overloaded profiles calculated using either the equilibrium-dispersive or POR model are in excellent agreement with the experimental profiles in all cases. Our results confirm that the competitive isotherm data derived from mixtures may suffice for a reasonably accurate prediction of the band profiles of all mixtures of the two components, provided their composition is close to 1/1. [source] Enantioseparation of novel chiral tetrahedral clusters on an amylose tris-(3,5-dimethylphenylcarbamate) chiral stationary phase by normal phase HPLCCHINESE JOURNAL OF CHEMISTRY, Issue 10 2004Wen-Zhi Li Abstract Amylose tris(3,5-dimethylphenylcamate) (ADMPC) coated on a kind of small particle silica gel was prepared. On this ADMPC chiral stationary phase (CSP), the direct enantiomeric separation of six novel chiral transition metal tetrahedral clusters has firstly been achieved using n -hexane as the mobile phase containing various alcohols as modifiers. The effect of mobile phase modifiers and the structural variation of the solutes on their retention factors (k) and resolutions (Rs) were investigated. The result suggests that not only the structure and concentration of alcohol in mobile phase, but also the structural differences in racemates can have a pronounced effect on enantiomeric separation. ADMPC-CSP is a suitable CSP for the optical resolution of chiral tetrahedral cluster by HPLC. [source] Theoretical reassessment of Whelk-O1 as an enantioselective receptor for 1-(4-halogeno-phenyl)-1-ethylamine derivativesCHIRALITY, Issue S1 2004Alberto Del Rio Abstract A combination of molecular mechanics and first principles calculations was used to explore the enantioselectivity of receptors, taking into account experimental data from the CHIRBASE database. Interactions between the Whelk-O1 HPLC chiral stationary phase with the complete series of 1-(4-halogeno-phenyl)-1-ethylamine derivative racemates were studied. The objective was to extract information from the interactions between the chiral Whelk-O1 stationary phase and the enantiomers, hence probing the origin of the enantioselective behavior. Calculations correctly reproduce the elution orders and reasonably describe the experimental enantioselectivities and retention factors. Different binding modes were observed for the first eluted enantiomer complexes, whereas the second eluted show only one prevalent diastereomeric binding fashion. Natural bond orbital (NBO) analysis was used on the global minima bound-complexes to quantify donor-acceptor interactions among chiral stationary phase and ligand moieties. Intermolecular hydrogen bonding was found to be the essential energetic interaction for all systems studied. CH-,, aromatic stacking and various charge transfer interactions were found to be smaller in magnitude but still important for the global enantioselective behavior. The three-point interaction model is discussed, pointing out the difficulty of its application for the qualitative prediction of elution orders (absolute configurations). Chirality 16:S1,S11, 2004. © 2004 Wiley-Liss, Inc. [source] | |||||||||||||