Supercritical Mobile Phase Research Articles

Mass spectrometry coupling of chip-based supercritical fluid chromatography enabled by make-up flow-assisted backpressure regulation

This work introduces a novel microfluidic backpressure pressure control developed for chip-based supercritical fluid chromatography (chipSFC). The presented on-chip pressure control mechanism involves the post-column addition of a viscous make-up stream, which enables pressure regulation within the range of 73 to 130 bar range. In contrast to approaches using mechanical backpressure regulators, this chip-based make-up-assisted pressure regulation offers a wear-free alternative that functions entirely through fluidic means and contributes minimally to extra column volume. It prevents phase separation of the supercritical mobile phase and, therefore, expands the analytical scope of chipSFC to detection systems with an ambient pressure interface. This was demonstrated by a proof-of-principle experiment, where a model mixture was separated within 30 s and detected using atmospheric pressure ionisation mass spectrometry.Graphical abstract

Gradient supercritical fluid chromatography coupled to mass spectrometry with a gradient flow of make-up solvent for enantioseparation of cathinones

In the past two decades, supercritical fluid chromatography has evolved from a niche application to a comprehensive technology and a fully-fledged alternative to conventional high-performance liquid chromatography.In this study, we have focused on chiral separation of synthetic cathinones in gradient supercritical fluid chromatography coupled to mass spectrometry using an inverse gradient of a make-up solvent. Synthetic cathinones possess an amphetamine-like effect and, therefore, are frequently being offered on the Internet as a replacement for illicit drugs. Cathinones are chiral compounds, however, they are usually marketed and used as racemic mixtures. Since the effect of individual enantiomers can significantly vary, there is a need for the development of enantioseparation methods enabling to study the biological effects of individual enantiomers. Since cathinones are basic molecules, they are easily protonated (positively charged) under weakly acidic mobile phase conditions, which is a typical feature of supercritical mobile phases with an alcohol as an organic modifier. The positively charged species represent ideal analytes for ion exchangers, such as chiral zwitterion ion exchangers Chiralpak ZWIX (+) and Chiralpak ZWIX (−), which possess a positively and negatively charged unit in the molecular structure of the selectors. The presence of the positive charge in the selector's structure, functioning as a counter-ion for the positively charged analytes, significantly reduces the required amount of a buffer, which is plausible for hyphenation of such a separation system with mass spectrometry. For mass spectrometry hyphenated to supercritical fluid chromatography, the use of a make-up solvent is required to avoid analyte precipitation when using a low concentration of an organic co-solvent (modifier) in the super-/subcritical mobile phase. Hereby, we introduce a unique approach, which is based on the gradient introduction of the make-up to the post-column effluent. Using this approach, it is possible to keep constant the overall amount of the organic solvent (modifier and make-up) introduced into the mass spectrometer when using a gradient of the organic modifier. We show that the developed gradient elution method facilitates the chiral separation of all employed analytes, while the mobile-phase gradient compensation by the inverse make-up gradient enables their detection with high signal intensities.

Advances in Profiling of Natural Products by Triple Detection Techniques Combined with Supercritical CO2 Mobile Phases and Sub 2-µm Stationary Phases

Advances in Profiling of Natural Products by Triple Detection Techniques Combined with Supercritical CO2 Mobile Phases and Sub 2-µm Stationary Phases

Advances in profiling of natural products by triple detection techniques combined with supercritical CO2 mobile phases and sub-2 µm stationary phases

Convergence chromatography (CC) is a separation technique that utilizes both super and sub-critical carbon dioxide (CO2) and sub-2 μm stationary phases to achieve unique selectivity, low solvent usage and high efficiencies. The use of supercritical fluid as a mobile phase provides higher diffusivity and lower viscosity than liquid mobile phases, thereby providing higher throughput and chromatographic efficiencies as compared to liquid chromatography.

Considerations on comprehensive and off‐line supercritical fluid chromatography×reversed‐phase liquid chromatography for the analysis of triacylglycerols in fish oil

The separation of the triacylglycerols in fish oil was performed by comprehensive and off-line supercritical fluid chromatography combined with RP-LC. The first dimension consisted of two serially coupled silver-ion (SI)-loaded columns operated with a supercritical mobile phase (supercritical fluid chromatography, SFC) in both the cases, whereas the second dimension was performed in non-aqueous RP mode (NARP-LC) on a 10-cm monolithic octadecyl silica (ODS) or a 45-cm long ODS column packed with 1.8 microm particles for the comprehensive and off-line separations, respectively. Despite the outstanding performance of the SI-SFC x NARP-LC interface, the high complexity of the sample rendered the online separation far from complete. The off-line approach gave much better separation mainly because of the higher peak capacity of the second-dimension column, but even in this case, the use of MS was mandatory to elucidate the different triacylglycerols in fish oil. The disadvantage of the off-line procedure was the long analysis time.

Extended Simulated Distillation by Capillary Supercritical Fluid Chromatography

Simulated distillation (simdist) based on separation using gas chromatography (GC) is widely spread in the petroleum industry for evaluation of fossil fuels as well as petroleum feeds and cuts produced by refining and conversion processes. Through a calibration curve relating the boiling point of normal paraffins to their elution temperature or retention time, simdist provides the hydrocarbon distribution of the sample (in weight percent) versus the boiling range of the fraction. The operating conditions used for simdist are tuned to be in agreement with preparative distillation that gives the True Boiling Point (TBP) curve. However, this technique is not compatible to heavy ends (FBP higher than 700°C) due to the limitation of volatility of the products and the lack of resistance of high molecular-weight hydrocarbons to cracking reactions. The interest of supercritical-fluid chromatography (SFC) is that high temperatures are not needed to elute heavy ends and that the mechanism for extending the upper limit of simdist depends only on sample solubility into the supercritical mobile phase. This paper presents recent advances in Simdist using SFC of heavy fractions. Compared to GC, SFC extends the range of this application up to nC126 hydrocarbons and makes possible to calibrate up to nC200. Thus, SFC could be the tool of choice for better determination of conversion in heavy petroleum-fraction processing.

Effect of system variables involved in packed column supercritical fluid chromatography of stavudine taken as model analyte using response surface methodology along with study of thermodynamic parameters

A multifactor optimization technique is successfully applied to study the effect of simultaneously varying the system variables on feasibility of stavudine analysis by packed column supercritical fluid chromatography (PC-SFC). The effect of simultaneously varying the pressure, temperature and modifier concentration was studied to optimize the method in order to obtain excellent chromatographic figures of merit. The method is based on isocratic elution using methanol-modified supercritical carbon dioxide as the mobile phase at the flow rate of 3.0 ml/min through a JASCO Finepak SIL-5, ODS [C 18 (5 μm, 25 cm × 4.6 mm, i.d.)] column support using photodiode array detection. The optimal conditions were determined with the aid of the response surface methodology using 3 3 factorial designs. From the response surface graphs optimum regions were selected to be +1, −1, and +1 for temperature (60 °C), pressure (20 MPa) and percent modifier concentration (17.81%, v/v), respectively. Linearity dynamic range was found to be in the range of 2.0–150.0 μg/ml with significantly high value of correlation coefficient. The method was validated for precision, robustness and recovery to assess the viability of the established method. The chromatographic limit of detection and quantitation were 0.80 and 1.50 μg/ml respectively. The method has been successfully used to analyze commercial dosage form to assess the chromatographic performance of SFC system which was found to be 99.91% ± 1.62. The present work briefs the thermodynamic applications of PC-SFC with an emphasis on the results of stavudine. The foremost of such applications is the determination of solute diffusion coefficient in supercritical mobile phase by Taylor–Aris peak broadening technique.

Effect of system variables involved in packed column SFC of nevirapine as model analyte using response surface methodology: Application to retention thermodynamics, solute transfer kinetic study and binary diffusion coefficient determination

A multifactor optimization technique is successfully applied to study the effect of simultaneously varying the system variables on feasibility of nevirapine analysis by packed column supercritical fluid chromatography (PC-SFC). The optimal conditions were determined with the aid of the response surface methodology using 3 3 factorial designs. The method is based on methanol-modified carbon dioxide as the mobile phase at flow rate of 3.0 ml/min with elution through a JASCO Finepak SIL-5, [C 18 (5-micron, 25 cm × 4.6 mm, i.d.)] column using photodiode array detection. The method has been successfully used to analyze commercial solid dosage form to assess the chromatographic performance of SFC system. The present work briefs the thermodynamic applications of PC-SFC with an emphasis on the results of nevirapine. The foremost of such applications is the determination of solute diffusion coefficient in supercritical mobile phase by Taylor–Aris peak broadening technique.

Supercritical Fluid Chromatographic Separation of Dimethylpolysiloxane Polymer

The use of compressed (dense) gases and supercritical fluids as chromatographic mobile phases in conjunction with liquid chromatographic (LC)-type packed columns was first reported by Klesper et al. in 1962. During its relatively short history, supercritical fluid chromatography (SFC) has become an attractive alternative to GC and LC in certain industrially important applications. SFC gives the advantage of high efficiency and allows the analysis of nonvolatile or thermally labile mixtures. The density of the mobile phase in SFC is about 200-500 times that in gas chromatography. The effect of shorter intermolecular distances and the resulting increase in molecular interactions is an enhanced solubilizing capability of the solvent towards various solutes. Compounds with much higher molecular weights than gas chromatography normally allows can therefore be chromatographed. However, the most commonly used mobile phases in SFC are all relatively non-polar fluids. Carbon dioxide, the most widely used fluid, is no more polar than hexane, even at high densities. Solute polarity should be between that of the stationary phase and the mobile phase in order to have a well behaved separation. Few real samples contain only nonpolar solutes, so a major objective of research in SFC has been directed toward increasing the range of solute polarity that can be handled by the technique. To bring the SFC technique into routine use, mobile phases that are more polar than the commonly used carbon dioxide are necessary. The solvent strength of supercritical CO2, even at high density, is not sufficient for the elution of polar solutes. Polar mobile phase such as NH3 exhibit useful properties, but a more practical way to extend the range of compounds separable by SFC is to use a mixed mobile phase. The solubility of the solute in the supercritical phase can be influenced considerably by adding modifiers to the mobile phase. The use of modifiers has been reported by Jentoft and Gouw and by Novotny et al. The latter group showed that adding 0.1% 2-propanol to pentane as the mobile phase decreases the observed partition coefficient (K) values for many polynuclear aromatic hydrocarbons by 20-35%. Thus, the addition of modifiers (generally organic solvents) to a supercritical mobile phase changes the polarity of the mobile phase and also leads to deactivation of the column packing material. In capillary SFC, most separations are made with pure CO2 because of its compatibility with an flame ionization detection (FID); except for formic acid and water, the addition of any common modifier precludes the use of FID. Modifiers are essential in packed-column SFC for the elution of polar compounds and are extensively used. Several workers have reported the influence of modifiers on peak shape, selectivity and retention time in capillary and packed-column SFC. A simple and effective way to add modifiers to a supercritical fluid mobile phase is to use a saturator column, which is usually a silica column saturated with polar alcohols. In this work, water was used as a polar modifier and a μporasil column as a saturator column. The μ-porasil column was inserted between the pump outlet and the injection valve. During the passage of the supercritical fluid mobile phase through the silica column, a polar modifier (water) can be dissolved in the pressurized supercritical fluid. Dimethylpolysiloxane polymer has been known as more polar polymer than polystyrene polymer. Dimethylpolysiloxane polymer has never been separated using water modified mobile phase. In this paper, using a μ-porasil column as a saturator column, excellent supercritical fluid chromatograms of dimethylpolysiloxane oligomers were obtained.

Correlation Equation for Retention Factor and Resolution of Ibuprofen in SFC

Supercritical fluid chromatography (SFC) was considered for separating racemic ibuprofen. The chromatographic column (3.9 x 150 mm) was packed with Kromasil® CHI-TBB, and the mobile phase was supercritical carbon dioxidewith modifier of IPA. The experimental variables were the content of IPA, and temperature and pressure of supercritical mobile phase. To determine the separation condition, the empirical equation of retention factor and resolution was proposed. In the case of retention factor, the empirical equation was in the form, k = a p + b/F + c (ρ/F) + d. The empirical equation for resolution was proposed as a linear form, R = a ρ + b F + c.

Supercritical argon as a mobile phase for the flame photometric detection of sulfur

The background emission properties of supercritical argon and supercritical carbon dioxide mobile phases in packed column supercritical fluid chromatography (pSFC) with flame photometric detection (FPD) were compared. As column flow rates were increased toward common values used in pSFC, the carbon dioxide background emission grew enormously. The resulting emission spectrum displayed dominant features at wavelengths between 325 and 525 nm, consisting of a complex series of overlapping molecular emission band systems partly attributed to CO* and CH*. By comparison, when using the same flow rates with a supercritical argon mobile phase, the background emission was identical to that of the FPD flame without column effluent. In terms of intensity, when using a column flow rate of 2 mL/min, supercritical carbon dioxide contributes a background emission in the FPD that is about 3 × 105 times larger than that produced by supercritical argon. This difference leads to an improvement of two orders of magnitude in the pSFC-FPD signal-to-noise ratio for sulfur when a supercritical argon mobile phase is used. Results indicate that supercritical argon could also be advantageous for the pSFC-FPD analysis of other elements.Key words: supercritical fluid chromatography, packed column, flame photometric detection, supercritical argon.

Separation of cis/ trans isomers of β-carotene by supercritical fluid chromatography

The aim of this work was to study and improve the separation of β-carotene cis/ trans isomers in SFC. Five different stationary phases have been tested, and their chromatographic behaviour has been determined. Unfortunately, because of the presence of additional isomers in the sample studied, none of the individual columns were able to isolate the four pure main isomers of β-carotene (9, 13, 15 cis and trans). The coupling of two different monomeric octadecyl columns was necessary to increase both selectivity and efficiency. The low viscosities of supercritical mobile phases allowed this column association. The influence of temperature, pressure, nature and modifier content were also investigated. Finally, with the optimum chromatographic conditions, six additional isomers have been separated from the four previous ones. One of them has been identified as the 9–9′ di- cis β-carotene.

Adsorptive separations using supercritical frontal analysis chromatography

AbstractMany food products and pharmuceuticals industries are looking to integrating super‐critical extraction and separations processes to fulfill processing needs due to limitations and regulations concerning the use of organic solvents. One such technology, which may affect these industries, is continuous supercritical adsorptive separation, a processing technology similar to the separation process of xylenes developed at UOP. Many products like pharmaceuticals and other human consumables could be processed more economically and with higher purity by supercritical fluid adsorption/desorption processes. Past studies, however, have focused on single‐component adsorption/desorption involving a supercritical mobile phase, whereas adsorptive separations involving super‐critical fluids are multicomponent systems. Accurate experimental techniques, which can determine both multicomponent solubilities in supercritical fluids and multicomponent adsorption isotherms in the presence of supercritical fluids, were developed to investigate the multicomponent supercritical adsorption phenomenon. A dynamic column model developed takes into account column dispersion as well as mass transfer and diffusive resistances. Experimental isotherm data incorporated into the model can predict the breakthrough profiles. An extension of pulse chromatography was used to identify the important hydrodynamic and transport parameters, and operating conditions are identified for optimal separations.

Modifier effects in open tubular capillary column supercritical fluid chromatography: Retention mechanisms from linear solvation energy relationships

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

Separation of Basic Drugs by Packed‐Column Supercritical Fluid Chromatography 3. Stimulants

A wide range of stimulants was rapidly and efficiently separated with symmetrical peak shapes with a tertiary supercritical mobile phase and a cyanopropyl liquid chromatography column. Compared with other drug families studied in this series, the stimulants exhibited a much broader range of retention. Several strong bases, including primary aliphatic amines, were easily eluted with a modified mobile phase. The stimulants were generally more retained than antipsychotic drugs. Compared with antidepressant drugs, a few stimulants behaved similarly, but most were more strongly retained. Modifier concentration proved to be the most effective means for changing both retention and selectivity. An over‐the‐counter eye drop solution (an aqueous buffer) was analyzed for tetrahydrozoline. The positive results with good peak shapes indicate that packed‐column supercritical fluid chromatography is compatible with at least small aqueous samples.

Band broadening during mobile phase change in unified chromatography (GC‐SFC)

AbstractBand broadening brought about by changing the mobile phase from the GC mode to the SFC mode in unified chromatography is discussed. Its origin is shown to result from several factors: the replaceable volume of the tubing and valves in front of and following the column; the volume, internal diameter, and film thickness of the open tubular column; the initial pressure of the supercritical mobile phase; and the column temperature. Improvements to restrict band broadening were achieved by optimizing the system and its operating conditions.

Solid-phase injector for open tubular column supercritical fluid chromatography

The design and use of a solid-phase injector for open tubular column supercritical fluid chromatography (SFC) are reported. For sample introduction, a liquid sample was loaded on a platinum wire. After the solvent had evaporated, the wire was inserted into, and sealed in, the injector. The chromatographic run was started by introducing the supercritical mobile phase into tie injector. Absolute and relative peak area reproducibilities, as well as retention time reproducibility, of the injection method were evaluated. The relative standard deviations (% RSDs) of absolute and relative peak areas were 2.0-4.5 and 0.4-1.8%, respectively, for both 1- and 2-μL injections

Optimal control of molecular resolution in supercritical-fluid chromatography with density programming

A technique which allows the a priori determination of optimal density programs for supercritical-fluid chromatography (SFC) is proposed. This new approach is meant to supersede heuristic methods for optimizing molecular resolution in SFC. The analysis is based upon optimal control theory applied to the treatment of molecular resolution in fixed beds with supercritical mobile phases. The optimal dynamic trajectories for the process operating conditions are found by solving the optimal control problem as posed here. A comparison with published data for a seven-component separation illustrates the concepts and shows that the method proposed has the potential for establishing optimal separation strategies in SFC.

Supercritical fluid-based cleanup technique for the separation of organochlorine pesticides from fats

A supercritical fluid cleanup technique for the separation of organochlorine pesticides from fats has been developed. The technique uses either an alumina column or a silica column with a supercritical mobile phase of carbon dioxide or methanol/carbon dioxide (2 mol % methanol), respectively. The lipid matrices were chicken fat with incurred residues of heptachlor epoxide, dieldrin, and endrin at low parts per million levels and lard spiked with lindane, heptachlor, hetachlor epoxide, dieldrin, en· drin, and o,p'·DDTat low parts per million levels. Recoveries (93-111 %) and precision (% RSD S 8.5) obtained with the supercritical fluid technique compare favorably with those obtained by conventional column cleanup methodology. In recent years, interest in the use of supercritical fluid extraction (SFE) as an analytical technique has increased considerably (Hawthorne, 1990). An interesting appli­ cation ofthis technology is in the field ofpesticide residue analyses. For instance, McNally and Wheeler (1988a,b) applied SFE to the analysis of sulfonylureas in soil, plant materials, and cell culture media. Engelhardt and Gross (1988) reported the analysis of lindane, aldrin, and p,p/. DDT in a spiked soil sample using SFE combined with supercritica1 fluid chromatography(SFC). Basicsolubility studies of pesticides in supercritical carbon dioxide have been conducted by Schafer and Baumann (1988, 1989). Recently, Lopez·Avila et al. (1989) reported the SFE of 17 organochlorine pesticides and 25 organophosphorus pesticides from sand using carbon dioxide or acetone­ modified carbon dioxide.

Optimization of derivatization parameters with 9-fluorenylmethyl chloroformate for amphetamine and catecholamine separation by supercritical fluid chromatography

The separation of amino compounds by supercritical fluid chromatography (SFC) is a difficult problem to solve, owing to the apolar nature of CO2. The derivatization of amino functions with the 9-fluorenylmethyl chloroformate (FMOC-Cl) allows to obtain apolar UV-absorbing compounds easily eluted with a supercritical mobile phase. Optimization of derivatization parameters allows us to analyze quantitatively amphetamines and catecholamines. These compounds can be separated in less than 5 min with a small addition of methanol as polar modifier. The total procedure takes no more than 15 min and can be automatized to gain time. As presented in this study, this method can be employed to physiological fluids as urine.