Reproducibility of sample separation using liquid or forced air convection thermostatted high performance capillary electrophoresis systems

Run-to-run sample separation reproducibility has been compared on two commercial high performance capillary electrophoresis units which differ in the mode by which the capillary temperature is thermostatted. Three standard analytes, differing dramatically in molecular character and size, were used for the analysis: benzoic acid, a 14 amino acid peptide from human chorionic gonadotropin, and ribonuclease A represent, respectively, small stable organic molecules, small peptides with little or no secondary structure, and proteins with secondary structure. These standards were evaluated with regard to reproducibility of migration time, peak area, and peak height. The analyses, performed in buffers of optimum pH for the separations, demonstrated that the liquid and forced air convection thermostatted systems both performed extremely well. The reproducibility, as judged by the percent coefficient of variance (% CV) of replicate analyses, was generally found to be less than 1 % (migration time); the reproducibility decreased in the order migration time > peak height > peak area. Whereas the absolute % CV values for MT_rel (migration relative to a standard) observed with the liquid thermostatted system were 2- to 4-fold lower than those observed with the forced air convection thermostatted system, there was little statistically significant difference between the two. As expected, the data indicated a reduction in reproducibility as the complexity of the analyte increased, perhaps as the result of an increased potential for wall interactions. Comparing separations in which low (≈?1 watt/meter [W/m] of capillary) and high (>5 W/m) Joule heat was generated by altering the sodium chloride content of the buffer revealed few statistically significant differences in the reproducibility obtained from the two systems. With these particular standard analytes and their respective buffer systems, there appears to be little difference between forced air convection and liquid thermostatting of the capillary.     

Estimation of binding constants between ristocetin and teicoplanin and peptides using on-column ligand derivatization coupled to affinity capillary electrophoresis

This work utilizes on-column ligand synthesis and affinity capillary electrophoresis (ACE) to determine binding constants (K_b) of 9-flourenylmethyloxy carbonyl (Fmoc)-amino acid derivatives to the glycopeptide antibiotics ristocetin (Rist) and teicoplanin (Teic). In this technique, two separate plugs of sample are injected on to the capillary column and electrophoresed. The initial sample plug contains a d-Ala-d-Ala terminus peptide and either one or two non-interacting standard(s). The second plug contains a Fmoc-amino acid-N-hydroxysuccinimide (NHS) ester. The electrophoresis is then carried out with an increasing concentration of Rist or Teic in the running buffer. Upon electrophoresis the initial d-Ala-d-Ala peptide reacts with the Fmoc-amino acid yielding a new Fmoc-amino acid-d-Ala-d-Ala peptide derivative. Continued electrophoresis results in the binding of Rist or Teic to the Fmoc-amino acid-d-Ala-d-Ala peptide derivatives. Analysis of the change in the relative migration time ratio (RMTR) or electrophoretic mobility () of the Fmoc-amino acid-d-Ala-d-Ala peptide derivatives relative to the non-interacting standards, as a function of the concentration of Rist and Teic, yields a value for K_b. These findings demonstrate the advantage of coupling on-column ligand synthesis to ACE for estimating binding parameters between antibiotics and ligands.Abbreviations Rist Ristocetin - Teic Teicoplanin - ACE Affinity capillary electrophoresis - RMTR Relative migration time ratio     

Multiple-injection affinity capillary electrophoresis to estimate binding constants of receptors to ligands

Multiple-injection affinity capillary electrophoresis (MIACE) is used to determine binding constants (K _b) between receptors and ligands using as model systems vancomycin and teicoplanin from Streptomyces orientalis and Actinoplanes teichomyceticus, respectively, and their binding to D-Ala-D-Ala peptides and carbonic anhydrase B (CAB. EC 4.2.1.1) and the binding of the latter to arylsulfonamides. A sample plug containing a non-interacting standard is first injected followed by multiple plugs of sample containing the receptor and then a final injection of sample containing a second standard. Between each injection of sample, a small plug of buffer is injected which contains an increasing concentration of ligand to effect separation between the multiple injections of sample. Electrophoresis is then carried out in an increasing concentration of ligand in the running buffer. Continued electrophoresis results in a shift in the migration time of the receptor in the sample plugs upon binding to their respective ligand. Analysis of the change in the relative migration time ratio (RMTR) or electrophoretic mobility (μ) of the resultant receptor–ligand complex relative to the non-interacting standards, as a function of the concentration of ligand yields a value for K _b. The MIACE technique is a modification in the ACE method that allows for the estimation of binding affinities between biological interactions on a timescale faster than that found for standard ACE. In addition sample volume requirements for the technique are reduced compared to traditional ACE assays. These findings demonstrate the advantage of using MIACE to estimate binding parameters between receptors and ligands.     

Interfacial tension coefficient from the retraction of ellipsoidal drops

A new method for the determination of the interfacial tension coefficient between two immiscible fluids is proposed. The method is particularly useful for the binary polymer blends. The deformed drop retraction method, DDRM, makes it possible to determine the dynamic interfacial tension coefficient, ν, from the time evolution of a distorted fluid drop toward its equilibrium form. Analysis of this interfacial tension-driven process led to a theoretical relation between the shape retraction rate and the system's geometrical and rheological characteristics. Measurements of either low viscosity model systems or high viscosity industrial polymer mixtures led to a good agreement with values obtained from the widely used breaking thread method. DDRM enables to measure ν in polymeric blends of commercial interest—the high viscosity systems that would be very difficult to characterize by other techniques. Furthermore, for the first time it is possible to follow the time dependence of the interfacial tension coefficient, thus unambiguously determine the dynamic and equilibrium values of ν₁₂. For example, in low density polyethylene blends with polystyrene, LDPE/PS, ν decreased with the polymer-polymer contact time, t_c, from ν = 6.9 mN/m at t_c = 12 min, to ν = 5.2 mN/m at t_c ≥ 75 min—the latter may represent the true thermodynamic equilibrium value, ν₁₂. However, it is not clear whether such a reduction is exclusively due to the thermodynamically driven migration of chain-ends, low molecular weight fractions and additives, or by the thermal degradation as well. The contact time dependence of ν explained some of the differences reported for the data obtained using different measurement techniques, viz. pendant drop, capillary breakup, or ellipsoid retraction techniques. © 1997 John Wiley & Sons, Inc. J Polym Sci B: Polym Phys 35 : 1393–1403, 1997     

Determination of the Enthalpies of Vaporization of Cyclic Organic Peroxides by Correlation of Changes in Gas Chromatographic Retention Times

Liquid and solid cyclic peroxides derived from aliphatic ketones are explosive materials so their enthalpies of vaporization and other thermodynamic or condensed-phase properties cannot be measured directly. In this work the enthalpies of vaporization of peroxides at 298.15 K were estimated simply from gas chromatographic retention times measured at different temperatures. The technique correlates changes in the retention times of compounds whose enthalpies of vaporization are known (called the reference series), with those of the compounds of interest. If t _R′ is the adjusted retention time (retention time of each compound minus the retention time of unretained diethyl ether, used as solvent) a plot of ln t _R′ against 1/T for each compound (reference compounds and cyclic peroxides) results in a straight line (r ² > 0.99 for all compounds). The enthalpy of transfer from solution to the vapor state (Δ_sol^g H _m) can be obtained by multiplying the slope by the gas constant (R). A second plot correlates the enthalpies of transfer from solution to the vapor state (Δ_sol^g H _m), as measured by gas–liquid chromatography (GLC), with enthalpies of vaporization of reference materials (Δ_vap H _m at 298.15 K) available in the literature. C₉–C₁₅ fatty acid methyl esters and hydrocarbons were used as reference compounds. The enthalpies of vaporization of the cyclic organic peroxides were calculated from the equation of the line obtained in this second correlation, the slope of which was Δ_vap H _m (at 298.15 K)/Δ^g _sol H _m. The experiments were performed under isothermal conditions with a DB-5 capillary column, flame-ionization detection (FID), and nitrogen as carrier gas. The column temperature was varied over a range of at least 30–70 K between 403 and 473 K, with chromatograms being acquired at 10 K intervals. Enthalpies of vaporization of cyclic organic peroxides are not available in the literature, and the values given in this paper, obtained by gas chromatography, are the first to be reported.     

UHF-AM1 studies on 1,4 and 1,5 intramolecular H-atom transfer reactions in alkenyl radicals

The transition states of intramolecular 1,4 and 1,5 H-atom transfers, from/to primary (p), secondary (s) or tertiary alkyl (t) and primary (p _a), secondary (s _a) or tertiary (t _a) allyl carbon atoms, have been studied at the level of the semiempirical quantum-chemical method AM1 with the UHF approximation. The activation and reaction enthalpies were calculated and compared with experimental data available in the literature and the calculated data obtained for analogous reactions in alkyl radicals. Correlations were found between the activation enthalpies and the dissociation enthalpies of the bonds broken and formed.     

Development of a CZE Method for the Determination of Olmesartan Medoxomil in Tablets

A CZE method was developed and validated for the analysis of Olmesartan medoxomil (OLMD) in tablets. The influences of pH, buffer concentration, applied voltage and capillary temperature on the migration time of OLMD were investigated. About 50 mM pH 6.5 phosphate buffer were used as background electrolyte. The optimum instrument parameters were found to be 30 °C temperature with 30 kV applied voltage and diode array detection was carried out at 210 nm. OLMD was hydrodynamically injected (P _inj  = 50 mbar, t _inj  = 3 s) and an internal standard, diflunisal (IS), was used to improve the precision and repeatability. Under these conditions, the migration time of OLMD was 2.32 min and the total analysis time was shorter than 5 min. Linearity range for the developed method was found to be 2.0–50.0 μg mL⁻¹ and the limit of detection was 0.5 μg mL⁻¹. The developed method was applied for the analysis of OLMD in pharmaceutical tablet formulations.

     

Identification of unknown diffusion and convection coefficients in ion transport problems from flux data: an analytical approach

This article presents an analytical approach for identification problems related to ion transport problems. In the first part of the study, relationship between the flux j_L : = (D(x)u_x(0, t)_x=0{\varphi_L := (D(x)u_x(0, t)_{x=0}} and the current response I(t){{\mathcal I}(t)} is analyzed for various models. It is shown that in pure diffusive linear model case the flux is proportional to the classical Cottrelian I_C(t){{\mathcal I}_C(t)}. Similar relationship is derived in the case of nonlinear model including diffusion and migration. These results suggest acceptability of the flux data as a measured output data in ion transport problems, instead of nonlocal additional condition in the form an integral of concentration function. In pure diffusive and diffusive-convective linear models cases, explicit analytical formulas between inputs (diffusion or/and convection coefficients) and output (measured flux data) are derived. The proposed analytical approach permits one to determine the unknown diffusion coefficient from a single flux data given at a fixed time t ₁ > 0, and unknown convection coefficient from a single flux data given at a fixed time t ₂ > t ₁ > 0. Linearized model of the nonlinear ion transport problem with variable diffusion and convection coefficients is analyzed. It is shown that the measured output (flux) data can not be given arbitrarily.     

Dynamic capillary diffusion system for monoterpene and sesquiterpene calibration: quantitative measurement and determination of physical properties

This article describes a method for preparation of low concentration gas standard mixtures of biogenic volatile organic compounds (BVOCs) emitted primarily by plants. A set of 10 plant volatiles including α-pinene, β-pinene, 3-carene, linalool, methyl salicylate, α-cedrene, β-caryophyllene, β-farnesene, aromadendrene and α-humulene was used in the study. Gas standard mixture of these compounds was generated using a capillary diffusion system (CDS). Diffusion coefficient (D) and saturation vapour pressure (p _s) data of these compounds were calculated from experimentally determined gas chromatographic retention indices (RI) and empirical relationships between D and p _s versus RI. A comparison of the calculated and measured concentrations of investigated compounds has proved that designed CDS can be successfully used for the proper quantification of BVOCs.     

Rapid determination of p<Emphasis Type="Italic">K</Emphasis><Subscript>a</Subscript> values of 20 amino acids by CZE with UV and capacitively coupled contactless conductivity detections

A rapid and universal capillary zone electrophoresis (CZE) method was developed to determine the dissociation constants (pK _a) of the 20 standard proteogenic amino acids. Since some amino acids are poorly detected by UV, capacitively coupled contactless conductivity detection (C⁴D) was used as an additional detection mode. The C⁴D coupling proved to be very successful on a conventional CE-UV instrument, neither inducing supplementary analyses nor instrument modification. In order to reduce the analysis time for pK _a determination, two strategies were applied: (i) a short-end injection to reduce the effective length, and (ii) a dynamic coating procedure to generate a large electroosmotic flow (EOF), even at pH values as low as 1.5. As a result, the analysis time per amino acid was less than 2 h, using 22 optimized buffers covering a pH range from 1.5 to 12.0 at a constant ionic strength of 50 mM. pK _a values were calculated using an appropriate mathematical model describing the relationship between effective mobility and pH. The obtained pK _a values were in accordance with the literature. Figure a UV (1) and C⁴D (2) detectors placed on-line on the CE capillary. b Curve of effective mobility as a function of pH for histidine     

Erratum: On the numerical stability of two widely used PLS algorithms

The above article (DOI: 10.1002/cem.1112) was published online on 14 February 2008. An error was subsequently identified: the captions for Figures 1 and 2 were omitted; they should read as follows: Figure 1. Orthogonality criterion (θ_A) for the octane data as a function of number of components (A) calculated using the standard PLS algorithm and SIMPLS. Figure 2. Orthogonality criterion (θ_A) for the wines data as a function of number of components (A) calculated using the standard PLS algorithm and SIMPLS.     

Cascades of energy among the vibrational levels of diatomic molecules trapped in a rare gas matrix: With application to 12C16O(ν) in Ar

Energy stored in vibrational level ν = 1 of several individual dipolar diatomic molecules AB which are trapped in a rare gas matrix M is automatically accumulated in a higher level ν > 1 of a single molecule AB. This remarkable cascade of energy upwards competes with a cascade of energy downwards. the radiative decay. The interplay of both cascades, first observed by Dubost and Charneau, is explained a simple model. The model incorporates three processes into a master equation for the relative populations P_ν(t) of levels ν: (a) migration of single quanta by resonance energy transfer, AB(1) + AB(0) ? AB(0) + AB(1); (b) phonon assisted excitation of upper levels, AB(1) + AB(ν) → AB(0) + AB(ν+1); and (c) radiative decay, AB(ν) → AB(ν-1). The model assumes that there is only one isotopic species AB which has a small but nonzero vibrational anharmonicity, that the temperature is low, T → 0 K, the concentration ratio ?_M/?_AB is large and that, initially, at time t = 0, a small fraction p₁ of molecules AB is excited to level ν = 1. The master equation has only two parameters, the radiative lifetime t_rad and k  2/[?_AB?₁k(1,1 → 0,2)t_rad], where k(1,1 → 0,2) is the reference rate constant of process (b). The master equation is solved in closed form for the Pν(t). For t_rad = 14 ms and k = 0.2, very satisfactory qualitative agreement is found for the theoretical P_ν(t) and the experimental time evolution of the relative population of vibrational levels of ¹²C¹⁶O in an argon matrix, for ?_M/?_AB = 2000 at T = 9 K. In agreement with experimental results it is concluded that the risetime of the fluorescence signals decreases whereas population inversion increases for decreasing values of ?_M/?AB. At long times, t > t_rad, any population inversion should disappear.     

Microcalorimetric Study of the Oscillating Extraction System

The power–time curves of the oscillating extraction system were determined at different temperatures for the extraction of hydrochloric acid and acetic acid with primary amine N₁₉₂₃ (R–CH(NH₂)–R¹), R, R ¹ represent alkyl of C_9–11 in chloroform using the titration microcalorimetric method. The apparent activation energy was calculated from the induction period (t _in), the first oscillation period (t ^p.1) and the second oscillation period (t _p.2). This revised version was published online in August 2006 with corrections to the Cover Date.     

Chain-length dependent termination in pulsed-laser polymerization, 2. On the prospects of determining the bimolecular termination constant kt from the second moment of the chain-length distribution

The correct (event weighted) average of k_t, 〈k_t〉, has been calculated for pseudostationary laser-induced polymerization for a kinetic scheme with chain-length dependent termination and compared to the average k̄_t obtained by formally solving for k_t the expression for the second moment of the chain-length distribution valid for chain-length independent termination (represented by the product of rate of polymerization ν_p and weight average degree of polymerization P̄_w). It is shown that there is a fair agreement between the two quantities. This may be used to recover experimentally the power-law governing the dependence of k_t on chain-length, especially its exponent.     

Estimation of <Superscript>235</Superscript>U concentration in some depleted uranium samples by high resolution gamma-ray spectrometry using 185 keV and 1001 keV gamma-energies of <Superscript>235</Superscript>U and <Superscript>234m</Superscript>Pa

The migration of ²³⁷Np in an undisturbed Chinese loess column was investigated by direct γ-ray method. The column was taken from a field test site and installed in a laboratory simulation hall. Radionuclide ²³⁷Np in the form of neptunium nitrate, mixed with quartz, was introduced into the column and covered with loess. Artificial rainfall was applied to the column for about 3 years and, the counting rates of ²³⁷Np in the column from 56 to 616 days at different vertical positions were detected with a γ-ray detection system. Based on the counting rates of ²³⁷Np in the simulation column at different vertical positions and the distance from the source layer, the relationship of the mass center of ²³⁷Np in the column at different experimental periods to the experimental time was established, C _m = 0.36 log(t)-2.75. Here C _m is the mass center of ²³⁷Np in the column, cm, and t is the experimental time in days. Based on this relationship, the mass center of ²³⁷Np for the 1,073-day experiment was predicted and compared to that obtained with the final destructive method. The good agreement between the prediction and the experimental values indicates that the direct γ-ray method could be used to predict the migration of strongly adsorbed radionuclides such as ²³⁷Np in environmental media with the help of laboratory simulation columns.     

Beiträge zur Chemie des Phosphors. 87. 1,2-Di-tert-butyl-3-iso-propyl-cyclotriphosphan,ein stabiles gemischt-substituiertes Cyclotriphosphan

Contributions to the Chemistry of Phosphorus. 87. 1,2-Di-tert-butyl-3-iso-propyl-cylclotriphosphane, a Stable Mixed-substituted Cyclotriphosphane The first kinetically stable mixed-substituted cyclotriphosphane, 1,2-di-tert-butyl-3-iso-propyl-cyclotriphosphane, (PBu^t)₂(PPrⁱ) ( 1 ), was synthesized by [2+1]-cyclocondensation of K(Bu^t)P–P(Bu^t)K with PrⁱPCl₂ in n-pentane. Mainly (PBu^t)₄ as well as mixed-substituted cyclotetra- and cyclopentaphosphanes are formed as by-products. 1 could be isolated in a pure state by high vacuum distillation and was thoroughly characterized. It forms two diastereomers, the more stable of which with a cis-standing tert-butyl and iso-propyl group can be stored without decomposition under inert conditions at room temperature for several days. Through thermolysis of 1 beside other alkylcyclophosphanes the mixed-substituted cyclotetraphosphanes (PBu^t)₂(PPrⁱ)₂ ( 2 ) and (PBu^t)₃(PPrⁱ) ( 3 ) are formed and their ³¹P NMR parameters are reported.     

Separation and determination of honokiol and magnolol in herbal medicines by flow injection-capillary electrophoresis

A simple, rapid, and accurate method for the separation and determination of honokiol and magnolol in Magnolia officinalis and related herbal medicines was developed by combination of flow injection (FI) and capillary zone electrophoresis (CZE). The analysis was carried out using an unmodified fused-silica capillary (50-μm I.D.; total length 7.5 cm; effective length 4.5 cm). A series of optimization steps afforded the following conditions: the sample solvent consisted of 150 mM NaOH and a running buffer composed of 10 mM sodium tetraborate/10 mM sodium dihydrogenphosphate (NaH₂PO₄) at pH 12 was applied for the separation of the analytes. The separation could be achieved within 5 min with a sample throughput rate of up to 28 h⁻¹. The repeatability (defined as the relative standard deviation, RSD) for honokiol and magnolol was 2.0% and 1.6% with peak area evaluation, 3.6% and 2.0% with peak height evaluation, and 2.0% and 1.4% with migration time evaluation, respectively. Regression equations revealed linear relationships (r = 0.9991–0.9998) between the peak area of each analyte and the concentration.     

Study of peak shape and efficiency in butyl acrylate-based monolithic columns for capillary electrochromatography

The study and modelling peak shape of in capillary electrochromatography (CEC), obtained using butyl acrylate (BA)-based monolithic columns, is described in this article. A modified-Gaussian model, which is a function of conventional experimental parameters: retention time (t_R), peak height (H₀) and standard deviation (σ₀) at the peak maximum, and left (A) and right (B) halfwidths, was used to describe the peaks of thiourea and several polyaromatic hydrocarbons compounds, which were eluted under several applied voltages. A mean relative error below 2% in the prediction of peak shape is obtained. Based on peak parameters, an easy and reliable estimation of global chromatographic performance, efficiency and peak capacity of BA-based monoliths was also considered. A comparison in terms of chromatographic performance of BA-based monoliths in CEC with CE mode and RPLC microparticulate columns was also performed.     

Gelation Studies: Comparison of the Critical Exponents Obtained by Dynamic Light Scattering and Rheology, 2

Summary: The sol–gel transition of two thermoreversible gelling mixtures made of xanthan gum and locust‐bean gum has been studied by using in situ, time‐resolved dynamic light scattering (DLS) and in situ rheology. A critical dynamical behavior was observed near the sol–gel transition, which was characterized by the presence of power‐law spectra over three and four decades in the time‐intensity correlation function g₂(t) − 1 ∼ t^−μ and over four and three decades in the oscillatory shear experiment G′(ω) ∼ G″(ω) ∼ ωⁿ. A comparison of the critical exponents obtained (μ₁ ≈ 0.36, μ₂ ≈ 0.32 and n₁ ≈ 0.62, n₂ ≈ 0.67) was made as a function of the dependence of the two mixing ratios according to the theory by Doi and Onuki. New experiments were also performed to compare the critical exponents on such a thermoreversible system.

Double‐logarithmic plot of the time‐intensity correlation functions g₂(t) − 1 versus the delay time, t, at a 90° scattering angle and at several temperatures of the mixture 1.