Electrochromatography in packed tubes using 1.5 to 50 μm silica gels and ODS bonded silica gels

Summary Electrochromatography (that is HPLC where the eluent is driven along the column by electro-osmosis using fields of up to 100 kV m⁻¹) promises plate efficiencies for HPLC which are comparable to those attained in capillary gas chromatography, but this requires that narrow-bore columns can be successfully packed with submicron particles. This paper demonstrates that we have now moved a considerable distance towards this goal. We show (1) that, following theory, there is no evidence of any reduction in electroosmotic velocity in columns packed with particles down to 1.5 μm diameter, (2) that reduced plate heights as low as unity are attainable for unretained solutes using both slurrypacked and drawn-packed columns 30 to 200 μm bore and up to 1 m long when packed with conventional 3 and 5 μm silica gels or with 1.5 μm impermeable silica spheres, (3) that columns driven electrically show higher plate efficiencies than identical columns driven by pressure, and (4) that 100,000 plate HPLC separations can be achieved in relatively short times of 30 minutes using in situ derivatised drawn packed capillaries containing 3 and 5 μm ODS-silica gels.     

Fast supercritical fluid chromatography of polymers using packed capillary columns

Summary Fast separations of perfluorinated polyethers and polymethylsiloxanes that are composed of 50–80 oligomers were demonstrated in packed capillary column supercritical fluid chromatography (SFC) using a carbon dioxide mobile phase. Separations were accomplished within 10 min using a 13 cm×250 μm i.d. column packed with 2 μm porous octadecyl bonded silica (ODS) particles. Effects of particle diameter of the packing material and pressure programming on separation were investigated, and packed column SFC was compared with open tubular column SFC. Results show that as the particle diameter was decreased from 5 to 3 to 2 μm and the column length was reduced from 85 to 43 to 13 cm, the separation time could be reduced from 70 to 20 to 10 min while still maintaining similar separation (resolution). Short columns packed with small porous particles are very suitable for fast SFC separations of polymers.     

Fast capillary electrophoresis-time-of-flight mass spectrometry using capillaries with inner diameters ranging from 75 to 5 μm

Fast electrophoretic separations in fused silica capillaries (CE) coupled to time-of-flight mass spectrometry (TOF-MS) are presented. CE separations of the model analytes (epinephrine, norepinephrine, dopamine, histidine, and isoproterenol) under conditions of high electric field strengths of up to 1.25 kV cm⁻¹ are completed in 20 s. Coupling of CE with TOF-MS is accomplished using a coaxial sheath liquid electrospray ionization interface. The influence of parameters inherent to the interface and their effects, including suction pressure and dilution, are discussed. In addition to standard capillaries of 75 and 50 μm inner diameter (ID), separations in capillaries with IDs of 25, 15, and 5 μm have been successfully applied to this setup. The analytical performance is compared over this range of capillary dimensions, and both advantages and disadvantages are discussed.     

Formation and characterization of mesostructured silica nanotubes

The multi-walled mesoporous silica nanotubes are prepared using cetyltrimethylammonium bromize (CTAB) as the surfactant micellar template and tetraethylorthosilicate (TEOS) as the silica precursor via a one-step wet chemical approach. The synthesized tubes are found to be double/triple walled and of amorphous nature. Their diameter and the length are about 100 nm to 1 μm and about 0.1–20 μm, respectively. The specific surface area approaches 1,488 m²/g. Based on the transmission electron microscopy analysis, it is inferred that the formation of the double/triple walled silica nanotubes is associated with the lamellar curling mechanism. A striking photoluminescence effect is detected in the mesostructured silica nanotubes. These nanotubes are expected to be a promising material for various applications such as gas storage, catalyst, or catalyst supports.     

Gas chromatography on 10 um silica particles

Summary Porous silica microparticles designed for modern liquid chromatography have proven effective in gas chromatography. Columns of 35–50 cm gave plate heights as low as 3.3 particle diameters and speeds of 2400 theoretical plates per second or 500 effective theoretical plates per second. Inlet pressures up to 70 atmospheres were required using hydrogen as carrier gas. The particles as received were too retentive for fast chromatography and gave asymmetric peaks. A coating of fluorosilicone oil overcame both problems. Other coatings were less effective. Bonded phases proved less satisfactory on both counts and also gave substantially less efficient columns and greater flow resistance. Column efficiency and flow resistance were sharply dependent on physical properties of the particles. The most efficient packing was clearly spherical particles of 5–10 μm diameter with narrow size distribution, pore diamters about 50 nm, BET surface areas of 25–50 m²/g and surfaces modified with trifluoropropyl silicone. A six-component hydrocarbon sample was separated in 33 s with a resolution of 4 for the most difficult pair and in 2.6 s with a minimal resolution. Performance was limited by end effects and by available pressure so that much better performance can be expected from longer columns and higher pressures.     

Effect of the pore sizes of nylon affinity membranes on γ-globulin adsorption

Summary l-Phenylalanine was immobilized on nylon membranes with two pore diameters (0.45 μm and 1 μm), by activation with 1,4-butanediol diglycidyl ether, and the effect of pore size on the affinity adsorption of γ-globulin studied by batch and kinetic methods. Experiment shows that adsorption on both affinity membranes obeys the Freundlich model. The accessible pore volume for adsorption of proteins on the membrane with 0.45 μm diameter pores is less than for that with 1 μm diameter pores. The adsorption capacity of affinity membranes with 1 μm diameter pores is 2.5-fold that of membranes with 0.45 μm diameter pores. Feed-rate has a larger effect on affinity adsorption on the membrane with 0.45 μm diameter pores than on that with 1 μm diameter pores. Small pores on the affinity membrane do not cause broadening of the elution peak. It is concluded that affinity interaction and separation occur mainly in the large pores, and small pore size does not favor improvement of adsorption capacity. γ-Globulin 85.1% pure can be obtained in one step from human plasma by use of the affinity membrane with 0.45 μm diameter pores.     

Determination of polar pesticides in soil by solid phase microextraction coupled to high-performance liquid chromatography-electrospray/mass spectrometry

The determination of carbamate and triazine pesticides from soil leachates and slurries was investigated using solid phase microextraction (SPME) coupled to high-performance liquid chromatography-electrospray/ mass spectrometry (HPLC-ESI/MS). SPME was carried out using fibres with a newly developed 50 μm Carbowax/ template coating which are suitable for relatively polar analytes. These fibers exhibit precisions better than 10% RSD, and are resistant against high contents of organic solvents during desorption. The technique shows a high sampling frequency resulting in an increasing sample throughput. Received: 17 December 1997 / Revised: 30 October 1998 / Accepted: 10 November 1998     

Controlling apatite microparticles formation by calcining electrospun sol–gel derived ultrafine silica fibers

Electrospun ultrafine silica fibers were calcined at 150–800 °C. The relation of calcination temperature to the ability to form biomimetic apatite in a simulated body fluid solution (SBF) was evaluated. The largest apatite particles, formed on non-calcined fibers after 1 week of soaking in SBF, were 10 μm in diameter, had a narrow size distribution (coefficient of variation 9%), and were similar to pearls on string. The particles size decreased with increasing calcination temperature below 250 °C and the particles formed on the fibers calcined at 250 °C were 4.5 μm in diameter. No particles were found on those calcined above 500 °C. By using a concentrated SBF at 1.5-times higher ionic concentrations than SBF, the size of apatite microparticles increased about 50%. The fibrous substrate covered with apatite particles was effective for osteoblastic differentiation of pre-osteoblastic cells.     

Synthesis of porous titaniaspheres for HPLC by polymerization-induced colloid aggregation (PICA) using <Emphasis Type="Italic">tert-n</Emphasis>-butyl titanate

Porous titanic microspheres with a very narrow particle size distribution (PSD) which are useful as chromatographic packing materials for high performance liquid chromatography (HPLC), were synthesized by an improved process of polymerization-induced colloid aggregation method using tert-n -butyl titanate. Porous titania particles obtained after polymer combustion and sintering of the aggregates are 3.5 μm in diameter with a surface area of 9.6 m² g⁻¹ and an average pore diameter of 18.9 nm. The particles are strong enough to withstand the high packing pressure for a HPLC column. The article is published in the original.     

Analysis of acrylamide in coffee and cocoa by isotope dilution liquid chromatography–tandem mass spectrometry

An accurate and precise method for the quantification of acrylamide using stable isotope dilution liquid chromatography–tandem mass spectrometry was developed and used to measure acrylamide in coffee and cocoa samples. The sample preparation involved extraction of the analyte and its internal standard, ¹³C₃-acrylamide, into water and subsequent defatting of the aqueous extract with dichloromethane. An aliquot of the resulting aqueous extract was then azeotropically dried under reduced pressure and subsequently purified using an aminopropyl-bonded silica cartridge. The purified extracts were then chromatographed on a 5-μm 2.1×150 mm Hypercarb column, the effluent of which was monitored for the analyte and its internal standard using positive-ion APCI-selected reaction monitoring. The intra-laboratory reproducibility of the method, expressed as a relative coefficient of variation (%, n=5), was determined at four levels of concentration (12.3, 42.3, 139.3 and 464.8 μg kg⁻¹) and was found to vary between 0.6–2.5%. The accuracy of the method was assessed using a reference sample of coffee. The average result obtained using our method differed from the assigned value of the reference material by less than 1%. An analysis of a cocoa sample revealed that the method is capable of precisely estimating acrylamide in challenging matrices down to a level of at least 12.3 μg kg⁻¹.     

Formation of Pores in Thermal Spray Coatings due to Incomplete Filling of Crevices in Patterned Surfaces

Molten particles in a thermal spray land on a rough surface, coalesce with each other and freeze to form a coating. Surface tension prevents liquid splats from completely filling crevices in the substrate, forming pores. An analytical model is developed to estimate the volume of such pores by calculating the equilibrium shape of a liquid meniscus pressing down on a surface asperity. Predictions from the model are compared with experimental results for the volume of voids formed under plasma sprayed yttria stabilised zirconia (YSZ) particles (average diameter 18 μm) landing with an average velocity of 250 m/s on patterned silicon surfaces that had vertical posts on them. The model predicted, to within an order-of-magnitude, the volume of voids on a surface in which the posts were tall (3 μm high) and closely spaced (1 μm apart), where pores were principally formed by incomplete filling of gaps.     

Control of flow rate in supercritical fluid chromatography

Summary Supercritical Fluid Chromatograph was constructed using two pumps and two restrictors. With this system in conjunction with the fused silica column packed with relatively large particles (40 μm), the flow rate was controlled both for isobaric and pressure programmed operations. Application of this system to the determination of molecular weight distribution of polystyrene oligomers was presented.     

Conversion of cellulose materials into nanostructured ceramics by biomineralization

Synthesis of hierarchically ordered silica materials having ordered wood cellular structures has been demonstrated through in-situ mineralization of wood by means of surfactant-directed mineralization in solutions of different pH. At low pH, silicic acid penetrates the buried interfaces of the wood cellular structure without clogging the pores to subsequently “molecularly paint” the interfaces thereby forming a positive replica following calcinations. At high pH, the hydrolyzed silica rapidly condenses to fill the open cells and pits within the structure resulting in a negative replica of the structure. Surfactant-templated mineralization in acid solutions leads to the formation of micelles that hexagonally pack at the wood interfaces preserving structural integrity while integrating hexagonally ordered nanoporosity into the structure of the cell walls following thermal treatment in air. The carbothermal reduction of mineralized wood with silica at high temperature produces biomorphic silicon carbide (SiC) materials, which are typical aggregations of β-SiC nanoparticles. To understand the roles of each component (lignin, crystalline cellulose, amorphous cellulose) comprising the natural biotemplates in the transformation to SiC rods, three different cellulose precursors including unbleached and bleached pulp, and cellulose nanocrystals have been utilized. Lignin in unbleached pulp blocked homogeneous penetration of silica into the pores between cellulose fibers resulting in non-uniform SiC fibers containing thick silica layers. Bleached pulp produced uniform SiC rods with camelback structures (80 nm in diameter; ∼50 μm in length), indicating that more silica infiltrates into the amorphous constituent of cellulose to form chunky rather than straight rod structures. The cellulose nanocrystal (CNXL) material produced clean and uniform SiC nanowires (70 nm in diameter; >100 μm in length) without the camelback structure.     

Simultaneous Determination of Triptolide and Tripdiolide in Extract of Tripterygium wilfordii Hook. f. by LC–APCI-MS

A novel method using liquid chromatography coupled with atmospheric pressure chemical ionization mass spectrometry in the negative selected ion monitoring mode has been developed and validated for rapid simultaneous determination of triptolide and tripdiolide in the extract of Tripterygium wilfordii Hook. f. The molecular ions m/z [M–H]⁻ 359 and 375 were selected for the quantification in selected ion monitoring mode for triptolide and tripdiolide. Standard calibration curve was linear over the concentration range of 0.12–24 and 0.15–30 μg mL⁻¹ for triptolide and tripdiolide. The relative standard deviations of intra- and inter-day were in the range of 4.7–9.9 and 8.9–12.6%. The average recoveries were between 96.4 and 104.6%. The limits of quantitation were 2.0 × 10⁻³ and 2.5 × 10⁻³ μg mL⁻¹ for triptolide and tripdiolide.     

Polybutadiene-coated zirconia packing materials in solvating gas chromatography using carbon dioxide as mobile phase

Summary In this paper, practical considerations of column efficiency, separation speed, thermal stability, and column polarity of capillary columns packed with polybutadiene-coated zirconia were investigated under solvating gas chromatography (SGC) conditions using carbon dioxide as mobile phase. When compared with results obtained from conventional porous octadecyl obtained from conventional porous octadecyl bonded silica (ODS) particles, PBD-zirconia particles produced greater change in mobile phase linear velocity with pressure than conventional ODS particles under the same conditions. The maximum plate number per second (N_t) obtained with a 30 cm PBD-zirconia column was approximately 1.5 times higher than that obtained with an ODS column at 100 °C. Therefore, the PBD-zirconia phase is more suitable for fast separations than conventional ODS particles in SGC. Maximum plate numbers per meter of 76,900 and 63,300 were obtained using a 57 cm×250 μm i.d. fused silica capillary column packed with 3 μm PBD-zirconia at 50 °C and 100 °C, respectively. The PBD-zirconia phase was stable at temperatures up to 320 °C under SGC conditions using carbon dioxide as mobile phase. Polarizable aromatic compounds and low molecular weight ketones and aldehydes were eluted with symmetrical peaks from a 10 cm column packed with 3 μm PBD-zirconia. Zirconia phases with greater inertness are required for the analysis of more polar compounds by SGC.     

Effect of pressure on solute capacity factor in HPLC using a non-porous stationary phase

Summary The pressure applied in liquid chromatography (LC) can influence the capacity factor of analytes. Using commercial equipment, column (33×4.6 mm) and 1.5 μm non-porous, tetradecyl (C₁₄) coated silica particles, a moderate but significant change in the capacity factor was observed with increasing pressure. Depending on the experimental conditions the relationship was found to be either linear or non-linear. In the latter case the retention increased at first, but later tended to decrease at still higher pressure. The results direct attention to the role of the pressure (and hence of flow rate and heat of friction) in determining capacity factors. Presented at Balaton Symposium on High Performance Separation Methods, Siófok, Hungary, September 1–3, 1999     

Determination of roxythromycin in blood serum by liquid chromatography with mass spectrometric detection

A procedure has been developed for the determination of a macrolide antibiotic roxythromycin (RX) in blood serum using HPLC with mass spectrometric detection using clarithromycin (CL) as the internal standard. RX and CL have been extracted from the samples by solid-phase extraction in a cartridge filled with a polar adsorbent, cyanopropylsilyl silica gel. The absolute recoveries of RX and CL are 89.6 and 92.5%, respectively. Chromatographic separation has been performed on a Nucleodur C₁₈ Isis column with the mobile phase composed as follows: water-methanol-acetonitrile-formic aid (499: 250: 250: 1 by volume). Registration has been performed in the mode of selected ion monitoring with m/z 837.7 (RX) and m/z 748.7 (CL). The analytical range for RX is 0.097–14.81 μg/mL, the quantification limit is 0.097 μg/mL, the detection limit is 0.03 μg/mL, and the intraday and interday relative standard deviation are 2–6 and 4–8% respectively. The procedure has been applied to the pharmacokinetic studies of the Rulid pharmaceutical preparation.     

Fast LC separation of a myoglobin digest: a case study using monolithic and particulate RP 18 silica capillary columns

A method was developed for the fast separation of a myoglobin digest using a monolithic RP 18 silica capillary column of 100 μm I.D. The results were compared with those obtained with a particulate RP 18 silica capillary column of 100 μm I.D. at a flow-rate between 0.6 and 1.2 μl/min. The digest was analyzed at the monolithic column at a flow-rate up to 2.8 μl/min. This high flow-rate could not be applied to the particulate column due to the high back-pressure. When the starting composition of the gradient was changed from 0 to 20% and a gradient steepness of 16%/min was used, the analysis time was less than 4 min. A positive Mascot identification score of 115 was achieved for the MS–MS data. When a lower gradient steepness was employed, the chromatographic resolution and the peak capacity did not increase for most compounds. The intraday repeatability for the retention time of the monolithic column was better than 1.5% at 2.8 μl/min and even less than 0.5% using a flow-rate of 0.6 or 1.0 μl/min. For the particulate column, it was between 0.5 and 1.4% for a flow-rate of 0.6 μl/min, probably due to the high column back-pressure. The interday reproducibility for the retention time of the monolithic column was less than 0.9% using a flow-rate of 1.0 μl/min.     

Evaluation of micro- versus nano-electrospray ionization for ambient pressure ion mobility spectrometry

The comparison of nanospray and microspray ionizations for detecting mixtures of compounds by ion mobility spectrometry has been investigated for sensitivity, ion transmission through a drift tube, and ion suppression effects when used as an ionization source for ambient pressure ion mobility spectrometry (IMS). Several articles have demonstrated that nano-electrospray ionization mass spectrometry (n-ESI-MS) has improved sensitivity, provides less background noise, and lower limits of detection than micro-electrospray ionization (μ-ESI) for IMS. Most importantly, data from n-ESI-MS is concentration-sensitive. Our laboratory previously published an article that observed a striking result when μ-ESI-IMS was investigated for a single compound in the positive ion mode. The data reported was mass-sensitive. In this new investigation, we have investigated mixtures, and experiments were designed to evaluate the effect of sensitivity, ion transmission and ion suppressions in μ-ESI-IMS and n-ESI-IMS. At an electrospray flow rate in the μL min⁻¹ range, compounds with higher proton affinities responded best while at the nanospray flow rates of nL min⁻¹, relative responses were more equal. This study observed that a decreased ESI flow rate resulted in a decreased ion signal. These trends demonstrated less sensitivity for ESI-IMS at reduced flow rates but suggest better quantification. At higher flow rates, relative ionization efficiencies were still uniform for all the components studied individually and in mixtures and sensitivity improved by about 78%. Concentration studies showed that at high concentrations, ion detection efficiencies were uniform at about 33% for all compounds studied individually and in mixtures. At low concentrations, the detection efficiency varied from 31% to 86%, depending on the proton affinity of the component in the mixture. Ion transmission through the IMS tube measured with a segmented Faraday detector that was incorporated into the IMS design indicated that most of the ion current for mixtures was transported through the IMS tube with a radius of less than 18 mm for both positive and negative ion modes.     

Purification of CIS-trimedlure isomers by high-performance liquid chromatography

Summary High-performance liquid chromatographic procedures were developed to make it possible to obtain the fourcis-trimedlure isomers (V, W, X, and Y) in pure form. Trimedlure-V and-Y were each readilt separated from the four-componentcis-trimedlure mixture through high-performance liquid chromatographic analysis on 5-μm and 3-μm silica, but trimedlure-W and-X were not adequately resolved. Chromatography of 5-μm silica of the mixtures obtained by epimerization of thetrans-trimedlure isomers, C and B₂, yielded the respective epimers, trimedlure-W and-X, in pure form.