Influence of the degree of coverage of C18-bonded stationary phases on the mass transfer mechanism and its kinetics

The influence of the degree of coverage of a silica surface with bonded C18 alkyl chains on the mass transfer mechanism in RPLC was investigated. Five packing materials were used, prepared with the same batch of silica particles (5 microm diameter, 90 A average pore size): one column was packed with the silica derivatized by trimethylchlorosilane (TMS) (C1, 3.92 micromol/m2), and the other four with the silica first derivatized with octadecyl-dimethyl-chlorosilane (C18, 0.42, 1.01, 2.03, and 3.15 micromol/m2), and then endcapped with TMS. A solution of methanol and water (25/75, v/v) was used as the mobile phase. The experimental HETP curves were acquired for each column by measuring the first moment and the second central moment of phenol and correcting them for the influence of the temperature increase due to the heat generated by the friction of the stream against the bed. The different kinetic parameters of the mass transfer in these packed chromatographic columns were identified (longitudinal diffusion, eddy diffusion, film mass transfer, and transparticle mass transfer) and quantified by fitting the experimental data to a new general HETP equation recently derived [F. Gritti, G. Guiochon, Anal. Chem., in press (AC-060203R).]. The agreement was excellent and allowed the comparison of the kinetic parameters among the six columns used. The highest column efficiency measured at conventional or fast flow rates (>0.5 ml/min) is obtained for the most retentive column, which has a surface coverage of 2.03 micromol/m2. The smallest HETP measured is as low as 10 microm, only twice the average particle diameter dp, due to the large contribution of surface diffusion (90%) to the particle effective diffusivity. However, no significant difference was observed between the efficiencies of the columns packed with C1 and C18 derivatized silica.     

Effects of the thermal heterogeneity of the column on chromatographic results

The influence of the thermal heterogeneity of HPLC columns on retention data was investigated. The retention factor of the retained compound phenol was measured at 24 increasing values of the flow rate, from 0.025 to 4.9 mL/min, on six different packing materials prepared with the same batch of silica particles (5 microm diameter, 90 A pore size). One column was packed with the neat silica particles, another with the silica endcapped with trimethylchlorosilane (TMS)(C(1), 3.92 micromol/m(2)), and the other four with silica first derivatized with octadecyl-dimethyl-chlorosilane (C(18), 0.42, 1.01, 2.03, and 3.15 micromol/m(2)), and second endcapped with TMS. Four different sources of heat contributing to raise the column temperature were considered: (1) the heat supplied by the hot high-pressure pump chamber to the solvent; (2) the adiabatic (dS=0) compression of the solvent in the high-pressure pump; (3) the isenthalpic (dH=0) decompression of the solvent during its migration along the porous chromatographic bed; and (4) the heat released by the friction of the solvent percolating through the column bed. The main contributions appear to be the heat supplied to the solvent by the HP pump and the friction heat. The average column temperature (ACT) was indirectly derived from the measurements of the first moment, mu(1), of phenol peak, of the column pressure drop, DeltaP, and of the retention factors of the phenol peak apices as a function of the flow rate applied. If the column is placed in a still-air bath at 298 K (and its temperature is not externally controlled), a longitudinal temperature gradient is established along the column and the average column temperature is about 6 K higher when this column is operated at 4.9 mL/min than when the flow rate is only 0.025 mL/min. If the column is placed in a heated air bath (temperature controlled at 316 or 338 K), the ACT changes by less than 3 K over the whole flow rate range applied.     

Influence of the pressure on the properties of chromatographic columns .III. retention volume of thiourea, hold-up volume, and compressibility of the C18-bonded layer

The influence of the average column pressure (ACP) on the elution volume of thiourea was measured on two RPLC columns, packed with Resolve-C18 (surface coverage 2.45 micromol/m2) and Symmetry-C18 (surface coverage 3.18 micromol/m2), and it was compared to that measured under the same conditions on an underivatized silica (Resolve). Five different methanol-water mixtures (20, 40, 60, 80 and 100% methanol, v/v) were used. Once corrected for the compressibility of the mobile phase, the data show that the elution volume of thiourea increases between 3 and 7% on the C18-bonded columns when the ACP increases from 50 to 350 bar, depending on the methanol content of the eluent. No such increase is observed on the underivatized Resolve silica column. This increase is too large to be ascribed to the compressibility of the stationary phase (silica + C18 bonded chains) which accounts for less than 5% of the variation of the retention factor. It is shown that the reason for this effect is of thermodynamic origin, the difference between the partial molar volume of the solute in the stationary and the mobile phase, Delta V, controlling the retention volume of thiourea. While Delta V is nearly constant for all mobile phase compositions on Resolve silica (with Delta V approximately equal to -4 mL/mol), on RPLC phases, it significantly increases with increasing methanol content, particularly above 60% methanol. It varies between -5 mL/mol and -17 mL/mol on Resolve-C18 and between -9 mL/mol and -25 mL/mol on Symmetry-C18. The difference in surface coverage between these two RP-HPLC stationary phases increases the values of Delta V by about 5 mL/mol.     

Feasibility of ultra high performance supercritical neat carbon dioxide chromatography at conventional pressures

The implementation of columns packed with sub-2 μm particles in supercritical fluid chromatography (SFC) is described using neat carbon dioxide as the mobile phase. A conventional supercritical fluid chromatograph was slightly modified to reduce extra column band broadening. Performances of a column packed with 1.8 μm C18-bonded silica particles in SFC using neat carbon dioxide as the mobile phase were compared with results obtained in ultra high performance liquid chromatography (UHPLC) using a dedicated chromatograph. As expected and usual in SFC, higher linear velocities than in UHPLC must be applied in order to reach optimal efficiency owing to higher diffusion coefficient of solutes in the mobile phase; similar numbers of theoretical plates were obtained with both techniques. Very fast separations of hydrocarbons are presented using two different alkyl-bonded silica columns.     

Adsorption mechanism in reversed-phase liquid chromatography. Effect of the surface coverage of a monomeric C18-silica stationary phase

The effect of the bonding density of the octadecyl chains onto the same silica on the adsorption and retention properties of low molecular weight compounds (phenol, caffeine, and sodium 2-naphthalene sulfonate) was investigated. The same mobile phase (methanol:water, 20:80, v/v) and temperature (T = 298 K) were applied and two duplicate columns (A and B) from each batch of packing material (neat silica, simply endcapped or C1 phase, 0.42, 1.01, 2.03, and 3.15 micromol/m2 of C18 alkyl chains) were tested. Adsorption data of the three compounds were acquired by frontal analysis (FA) and the adsorption energy distributions (AEDs) were calculated using the expectation-maximization method. Results confirmed earlier findings in linear chromatography of a retention maximum at an intermediate bonding density. From a general point of view, the saturation capacity of the adsorbent tends to decrease with increasing bonding density, due to the vanishing space intercalated between the C18 bonded chains and to the decrease of the specific surface area of the stationary phase. The equilibrium constants are maximum for an intermediary bonding density (approximately 2 micromol/m2). An enthalpy-entropy compensation was found for the thermodynamic parameters of the isotherm data. Weak equilibrium constants (small deltaH) and high saturation capacities (large deltaS) were observed at low bonding densities, higher equilibrium constants and lower saturation capacities at high bonding densities, the combinations leading to similar apparent retention in RPLC. The use of a low surface coverage column is recommended for preparative purposes.     

Effect of the surface coverage of endcapped C18-silica on the excess adsorption isotherms of commonly used organic solvents from water in reversed phase liquid chromatography

The excess adsorption isotherms of methanol, ethanol, 2-propanol, acetonitrile, and tetrahydrofuran from water were measured on five different silica-based packing materials by the minor disturbance method. These materials were prepared with the same lot of 5-microm particles (average pore size 90 A), all endcapped with trimethylchlorosilane (TMS), and bonded to octadecyl chains with different surface coverages (0, 0.42, 1.01, 2.03, and 3.15 micromol/m2). The relative adsorption of one eluent by respect to a second one informs on the heterogeneity of the material (alkyl-bonded and bare silica regions) and on the accessibility of the unreacted silanol groups to the mobile phase. It is shown that the total surface area of the adsorbent decreases with increasing degree of surface coverage with octadecyl chains and that the relative surface area of the regions occupied by accessible silanol groups to the regions occupied by alkyl-bonded groups decreases. For the five columns, an average of 10% of the adsorbent surface area is covered of bare silica accessible to the liquid phase, with a minimum of 5% with tetrahydrofuran and a maximum of 12% with ethanol or 2-propanol. Increasing the surface coverage by the C18 chains causes a significant increase of the attraction potential of the hydrophobic surface toward the organic solvent. This result is confirmed by the increase of the number of adsorbate monolayers with increasing bonding density of the octadecyl chains. This number is twice larger for the 315C18 column than for the C1 column.     

A chromatographic estimate of the degree of surface heterogeneity of reversed-phase liquid chromatography packing materials. II-Endcapped monomeric C18-bonded stationary phase

In a previous report, the heterogeneity of a non-endcapped C30-bonded stationary phase was investigated, based on the results of the measurements of the adsorption isotherms of two neutral compounds (phenol and caffeine) and two ionizable compounds (sodium naphthalene sulfonate and propranololium chloride) by frontal analysis (FA). The same method is applied here for the characterization of the surface heterogeneity of two new brands of endcapped C18-bonded stationary phases (Gemini and Sunfire). The adsorption isotherms of the same four chemicals were measured by FA and the results confirmed by the independent calculation of the adsorption energy distribution (AED), using the expectation-maximization (EM) method. The effect of the length of the bonded alkyl chain was investigated. Shorter alkyl-bonded-chains (C18 versus C30) and the end-capping of the silica surface contribute to decrease the surface heterogeneity under the same experimental conditions (30% methanol, 25 mM NaCl). The AEDs of phenol and caffeine are bimodal with the C18-bonded columns while they are trimodal and quadrimodal, respectively, with a non-endcapped C30-bonded column. The "supersites" (adsorption energy > 20 kJ/mol) found on the C30-Prontosil column and attributed to a cation exchange mechanism completely disappear on the C18-Gemini and C18-Sunfire, probably because the end-capping of the silica surface eliminates most if not all the ionic interactions.     

Radial heterogeneity of some analytical columns used in high-performance liquid chromatography

An on-column electrochemical microdetector was used to determine accurately the radial distribution of the mobile phase velocity and of the column efficiency at the exit of three common analytical columns, namely a 100 mm × 4.6 mm C18 bonded silica-based monolithic column, a 150 mm × 4.6 mm column packed with 2.7 μm porous shell particles of C18 bonded silica (HALO), and a 150 mm × 4.6 mm column packed with 3 μm fully porous C18 bonded silica particles (LUNA). The results obtained demonstrate that all three columns are not radially homogeneous. In all three cases, the efficiency was found to be lower in the wall region of the column than in its core region (the central core with a radius of 1/3 the column inner radius). The decrease in local efficiency from the core to the wall regions was lower in the case of the monolith (ca. 25%) than in that of the two particle-packed columns (ca. 35–50%). The mobile phase velocity was found to be ca. 1.5% higher in the wall than in the core region of the monolithic column while, in contrast, it was ca. 2.5–4.0% lower in the wall region for the two particle-packed columns.     

Physical origin of peak tailing on C18-bonded silica in reversed-phase liquid chromatography

Single component isotherm data of caffeine and phenol were acquired on two different stationary phases for RPLC, using a methanol/water solution (25%, v/v, methanol) as the mobile phase. The columns were the non-endcapped Waters Resolve-C18, and the Waters XTerra MS C18. Both columns exhibit similar C18 -chain densities (2.45 and 2.50 micromol/m2) and differ essentially by the nature of the underivatized solid support (a conventional, highly polar silica made from water glass, hence containing metal impurities, versus a silica-methylsilane hybrid surface with a lower density of less acidic free silanols). Thirty-two adsorption data points were acquired by FA, for caffeine, between 10(-3) and 24 g/l, a dynamic range of 24,000. Twenty-eigth adsorption data points were acquired for phenol, from 0.025 to 75 g/l, a dynamic range of 3000. The expectation-maximization procedure was used to derive the affinity energy distribution (AED) from the raw FA data points, assuming a local Langmuir isotherm. For caffeine, the AEDs converge to a bimodal and a quadrimodal distribution on XTerra MS-C18 and Resolve-C18, respectively. The values of the saturation capacity (q(s,1) approximately equal to 0.80 mol/l and q(s,2) approximately equal to 0.10 mol/l) and the adsorption constant (b1 approximately equal to 3.11/mol and b2 approximately equal to 29.1 l/mol) measured on the two columns for the lowest two energy modes 1 and 2, are comparable. These data are consistent with those previously measured on an endcapped Kromasil-C18 in a 30/70 (v/v), methanol/water solution (q(s,1) = 0.9 mol/l and q(s,2) = 0.10 mol/l, b1 = 2.4 l/mol and b2 = 16.1 l/mol). The presence of two higher energy modes on the Waters Resolve-C18 column (q(s,3) approximately equal to 0.013 mol/l and q(s,4) approximately equal to 2.6 10(-4) mol/l, b3 approximately equal to 252 l/mol and b4 = 13,200 l/mol) and the strong peak tailing of caffeine are explained by the existence of adsorption sites buried inside the C18-bonded layer. It is demonstrated that strong interactions between caffeine and the water protected bare silica surface cannot explain these high-energy sites because the retention of caffeine on an underivatized Resolve silica column is almost zero. Possible hydrogen-bond interactions between caffeine and the non-protected isolated silanol groups remaining after synthesis amidst the C18-chain network cannot explain these high energy interactions because, then, the smaller phenol molecule should exhibit similarly strong interactions with these isolated silanols on the same Resolve-C18 column and, yet, the consequences of such interactions are not observed. These sites are more consistent with the heterogeneity of the local structure of the C18-bonded layer. Regarding the adsorption of phenol, no matter whether the column is endcapped or not, its molecular interactions with the bare silica were negligible. For both columns, the best adsorption isotherm was the Bilangmuir model (with q(s,1) approximately equal to 2 and q(s,2) approximately equal to 0.67 mol/l, b1 0.61 and b2 approximately equal to 10.3 l/mol). These parameters are consistent with those measured previously on an endcapped Kromasil-C18 column under the same conditions (q(s,1) = 1.5 and q(s,2) = 0.71 mol/l, b1 = 1.4 l/mol and b2 = 11.3 l/mol). As for caffeine, the high-energy sites are definitely located within the C18-bonded layer, not on the bare surface of the adsorbent.     

Effect of the density of the C(18) surface coverage on the adsorption mechanism of a cationic compound and on the silanol activity of the stationary phase in reversed phase liquid chromatography

RPLC columns with different surface coverages (a C(1) endcapped column with a bonding density of 3.92 micromol/m(2) and four C(18)-bonded, endcapped columns, with octadecyl chain densities of 0.42, 1.01, 2.03, and 3.15 micromol/m(2)) were used to investigate the effects of the density of the surface coverage of RPLC columns on the adsorption mechanism of a cationic compound, amitriptyline chloride, and on the silanol activity of these columns. The mobile phases used were acetonitrile-water (30/70, v/v) solutions, buffered at either pH 2.7 or pH 6.9. At pH 2.7, the residual silanol groups are not ionized. At pH 6.9, some of these groups are ionized and these surface anions can strongly interact with the cationic compound. The adsorption isotherms were measured by frontal analysis (FA) at pH 2.7 and by frontal analysis by characteristic points (FACP) at pH 6.9, because the very high retention observed at neutral pH made FA measurements excessively long and poorly accurate. The adsorption energy distributions (AEDs) were calculated when possible, according to the expectation-maximization (EM) algorithm. A bimodal and a trimodal energy distribution were found for all the columns at pH 2.7 and 6.9, respectively. The third site measured at pH 6.9 was attributed to the strong ion-exchange interactions between the ionized silanol groups and the amitriptylinium cation. The contribution of the ionized silanol groups to the overall retention is maximum for the phases with intermediary bonding densities (1.01 and 2.03 micromol/m(2)). The peak tailing is most pronounced for the lowest (C(1) column) and the highest (3.15 micromol/m(2)) surface coverages.     

Monolithic silica rod columns for high-efficiency reversed-phase liquid chromatography

Chromatographic properties of a new type of monolithic silica rod columns were examined. Silica rod columns employed for the study were prepared from tetramethoxysilane, modified with octadecylsilyl moieties, and encased in a stainless-steel protective column with two polymer layers between the silica and the stainless-steel tubing. A 25 cm column provided up to 45,000 theoretical plates for aromatic hydrocarbons, or a minimum plate height of about 5.5 μm, at optimum linear velocity of ca. 2.3 mm/s and back pressure of 7.5 MPa in an acetonitrile-water (80/20, v/v) mobile phase at 40°C. The permeability of the column was similar to that of a column packed with 5 μm particles, with K(F) about 2.4×10(-14) m(2) (based on the superficial linear velocity of the mobile phase), while the plate height value equivalent to that of a column packed with 2.5 μm particles. Generation of 80,000-120,000 theoretical plates was feasible with back pressure below 30 MPa by employing two or three 25 cm columns connected in series. The use of the long columns enabled facile generation of large numbers of theoretical plates in comparison with conventional monolithic silica columns or particulate columns. Kinetic plot analysis indicates that the monolithic columns operated at 30 MPa can provide faster separations than a column packed with totally porous 3-μm particles operated at 40 MPa in a range where the number of theoretical plates (N) is greater than 50,000.     

Silica particles encapsulated poly(styrene-divinylbenzene) monolithic stationary phases for micro-high performance liquid chromatography

In the paper we demonstrate a new approach for the preparation and application of continuous silica bed columns that involve encapsulation (entrapment) of functionalized silica microparticles, which can be used as packing material in micro high performance liquid chromatography (micro-HPLC) and capillary electrochromatography (CEC). Like traditional packed columns, these capillaries possess characterized silica particles that offer high phase ratio and narrow pore size distribution leading to high retention and separation efficiency, respectively. More importantly, immobilization of the microparticles stabilizes the separation bed and eliminates the need for retaining frits. The developed capillary columns were fabricated in exactly the same way as a packed capillary column (slurry packing) but with an additional entrapment step. This immobilization of the packed bed was achieved by in situ polymerization of styrene and divinylbenzene in presence of decanol as a porogen and azobisisobutyronitrile as thermal initiator. Silica particles with different particle sizes and pore sizes ranging from 60 to 4000 A were studied. In addition different modified silica was used, including C-18 reversed phase, anion exchange and chiral stationary phases. Efficient separation of polyphenolic compounds, peptides, proteins and even DNA mutation were achieved using the developed technique depending on the properties of the silica particles used (particles pore size). For example, using 3 microm ProntoSIL C-18 particles with 300 A pore size, separation efficiencies in the range of 120,000-200,000 plates/m were obtained for protein separation, in a 6 cm x 200 microm i.d. capillary column. Using encapsulated silica C-18 with 1000 A pore size, separation of DNA homo and hetero duplexes were achieved under denaturing HPLC conditions for mutation detection. In addition, nucleotides were separated using anion exchange material encapsulated with poly(styrene-divinylbenzene) (PS/DVB), which indicated that the chromatographic properties of the silica packing material were still active after polymerization. The prepared capillary columns were found to be stable and could easily be operated continuously up to a pressure of 350 bar without column damage and capillary can be cut to any desired length.     

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.     

Monolithic silica columns with various skeleton sizes and through-pore sizes for capillary liquid chromatography

Reduction of through-pore size and skeleton size of a monolithic silica column was attempted to provide high separation efficiency in a short time. Monolithic silica columns were prepared to have various sizes of skeletons (approximately 1-2 microm) and through-pores (approximately 2-8 microm) in a fused-silica capillary (50-200 microm I.D.). The columns were evaluated in HPLC after derivatization to C18 phase. It was possible to prepare monolithic silica structures in capillaries of up to 200 microm I.D. from a mixture of tetramethoxysilane and methyltrimethoxysilane. As expected, a monolithic silica column with smaller domain size showed higher column efficiency and higher pressure drop. High external porosity (> 80%) and large through-pores resulted in high permeability (K = 8 x 10(-14) -1.3 x 10(-12) m2) that was 2-30 times higher than that of a column packed with 5-mirom silica particles. The monolithic silica columns prepared in capillaries produced a plate height of about 8-12 microm with an 80% aqueous acetonitrile mobile phase at a linear velocity of 1 mm/s. Separation impedance, E, was found to be as low as 100 under optimum conditions, a value about an order of magnitude lower than reported for conventional columns packed with 5-microm particles. Although a column with smaller domain size generally resulted in higher separation impedance and the lower total performance, the monolithic silica columns showed performance beyond the limit of conventional particle-packed columns under pressure-driven conditions.     

Practical comparison of LC columns packed with different superficially porous particles for the separation of small molecules and medium size natural products

Commercial C(18) columns packed with superficially porous particles of different sizes and shell thicknesses (Ascentis Express, Kinetex, and Poroshell 120) or sub-2-μm totally porous particles (Acquity BEH) were systematically compared using a small molecule mixture and a complex natural product mixture as text probes. Significant efficiency loss was observed on 2.1-mm id columns even with a low dispersion ultra-high pressure liquid chromatography system. The Kinetex 4.6-mm id column packed with 2.6-μm particles exhibited the best overall efficiency for small molecule separations and the Poroshell 120 column showed better performance for mid-size natural product analytes. The Kinetex 2.1-mm id column packed with 1.7-μm particles did not deliver the expected performance and the possible reasons besides extra column effect have been proved to be frictional heating effect and poor column packing quality. Different column retentivities and selectivities have been observed on the four C(18) columns of different brands for the natural product separation. Column batch-to-batch variability that has been previously observed on the Ascentis Express column was also observed on the Kinetex and Poroshell 120 column.     

Fast, accurate, and convenient analysis of bed densities for columns packed with fine reversed-phase particles

We determine the interparticle porosities of commercially available, analytical, reversed-phase HPLC columns by Donnan exclusion of a small, unretained, co-ionic tracer (nitrate ions). The columns contained packings of C(18)-modified, endcapped, silica particles, which differed in their nominal particle diameters (1.8-5 μm) and construction (fully porous or core-shell). Experiments were carried out by monitoring the elution volumes of nitrate samples in a mobile phase of acetonitrile/water 80:20 v/v at increasing concentrations of Tris-HCl buffer (pH 8.1) from 0.01 to 60 mM. At low buffer concentrations, nitrate ions are completely electrostatically excluded from the intraparticle mesopore space, which is reflected by a plateau region in the elution curves. The elution volume in the plateau region equals the interparticle void volume. Clearly defined plateau regions were observed for all columns, even those densely packed with core-shell and sub-2 μm particles, enabling the accurate determination of interparticle porosities to three decimal places in a fast and convenient way.     

Ultra high pressure liquid chromatography. Column permeability and changes of the eluent properties

The behavior of four similar liquid chromatography columns (2.1mm i.d. x 30, 50, 100, and 150 mm, all packed with fine particles, average d(p) approximately 1.7 microm, of bridged ethylsiloxane/silica hybrid-C(18), named BEH-C(18)) was studied in wide ranges of temperature and pressure. The pressure and the temperature dependencies of the viscosity and the density of the eluent (pure acetonitrile) along the columns were also derived, using the column permeabilities and applying the Kozeny-Carman and the heat balance equations. The heat lost through the external surface area of the chromatographic column was directly derived from the wall temperature of the stainless steel tube measured with a precision of +/-0.2 degrees C in still air and +/-0.1 degrees C in the oven compartment. The variations of the density and viscosity of pure acetonitrile as a function of the temperature and pressure was derived from empirical correlations based on precise experimental data acquired between 298 and 373 K and at pressures up to 1.5 kbar. The measurements were made with the Acquity UPLC chromatograph that can deliver a maximum flow rate of 2 mL/min and apply a maximum column inlet pressure of 1038 bar. The average Kozeny-Carman permeability constant of the columns was 144+/-3.5%. The temperature hence the viscosity and the density profiles of the eluent along the column deviate significantly from linear behavior under high-pressure gradients. For a 1000 bar pressure drop, we measured DeltaT=25-30 K, (Deltaeta/eta) approximately 100%, and (Deltarho/rho) approximately 10%. These results show that the radial temperature profiles are never fully developed within 1% for any of the columns, even under still-air conditions. This represents a practical advantage regarding the apparent column efficiency at high flow rates, since the impact of the differential analyte velocity between the column center and the column wall is not maximum. The interpretation of the peak profiles recorded in UPLC is discussed.     

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.     

Separation of 2-naphthol atropisomers on cyclofructan-based chiral stationary phases

A set of fifteen 2-naphthol-derived atropisomers were evaluated on three different cyclofructan-based chiral stationary phases (CSP). The cyclofructan CSPs were a dimethylphenyl-derivatized cyclofructan 7 (CF7-DMP), a isopropyl (CF6-P) and a R-naphthylethyl cyclofructan 6 (CF6-RN) derivative, all bonded to 5-µm spherical fully porous silica particles packed into three 25?cm?×?4.6?mm columns (commercially available as Larihc columns). The 15 atropisomers were analyzed in the normal-phase mode with heptane/alcohol mobile phases. The CF7-DMP column was the most effective partially or fully separating 14 of the 15 compounds (93%). All 15 compounds could be separated by at least one cyclofructan column. Five compounds could be partially or fully separated by all three CSPs. The effect of ethanol, 2-propanol and butanol as 5 and 10% v/v polar modifiers in heptane was studied. A prototype 15?cm?×?4.6?mm column packed with superficially porous 2.7?µm CF6-P bonded particles was tested with the same set of compounds and a standard HPLC system. The increased efficiency and solvent saving were significant.     

Monolithic Silica Columns for HPLC,Micro‐HPLC,and CEC

Two types of monolithic silica columns derivatized to form an ODS phase, one prepared in a fused silica capillary (SR‐FS) and the other prepared in a mold and clad with an engineering plastic (poly‐ether‐ether‐ketone) (SR‐PEEK), were evaluated. The column efficiency and pressure drop were compared with those of a column packed with 5‐μm ODS‐silica particles and of an ODS‐silica monolith prepared in a mold and wrapped with PTFE tubing (SR‐PTFE). SR‐FS gave a lower pressure drop than a column packed with 5‐μm particles by a factor of 20, and a plate height of 20 μm at a linear velocity below 1 mm/s. SR‐PEEK showed higher flow‐resistance than the other monolithic silica columns, but they still showed a minimum plate height of 8–10 μm and a lower pressure drop than popular commercial columns packed with 5‐μm particles. The evaluation of SR‐FS columns in a CEC mode showed much higher efficiency than in a pressure‐driven mode.