Use of various filters for generating modified supercritical fluid mobile phases

Summary Adding various components to supercritical carbon dioxide in supercritical fluid chromatography can extend or significantly alter the solvating properties. Polar samples which are difficult to analyze with pure supercritical CO₂ because of their high polarity can be separated by addidng polar modifiers. In this paper, a new mixing method using an HPLC filter for adding polar modifier to CO₂ is described. Although several filters were tried, only one could keep the amount of modifier in the mobile phase constant for a long time. The amount of water or methanol dissolved in supercritical CO₂ was measured by an amperometric microsensor made of a thin film of perfluorosulfonate ionomer (PFSI).     

New device for controlling the amount of methanol in carbon dioxide mobile phase for supercritical fluid chromatography

A new device to accurately deliver a small amount of methanol into supercritical carbon dioxide fluid is described. Carbon dioxide, the most widely used mobile phase in supercritical fluid chromatography, is a relatively non-polar fluid, and hence the addition of a small amount of methanol could change the solvent strength of the mobile phase. In this work, supercritical CO₂ and methanol are delivered from the pump to a 100-μl mixing chamber in which a small magnetic bar is rotating. After passing through the mixing chamber, supercritical CO₂ is changed to a new mobile phase with different polarity. The modified mobile phase was successfully used for the separations of polar compounds and polyaromatic hydrocarbons (PAHs).     

Application of a gas-liquid entraining rotor to supercritical fluid extraction : Removal of iron(III) from water

Supercritical fluid extraction (SFE) of aqueous solutions is often limited by poor mass transport. The performance of a new gas-liquid entraining device was investigated to improve mass transport and thereby increase extraction efficiency. As a test system, iron(III) was extracted from water with a β-diketone chelating agent (HL) and supercritical fluid carbon dioxide. Metal β-diketonate complexes with sufficient solubility in supercritical fluid CO₂ are often poorly extracted from aqueous solutions due to limited mass transport between the water-soluble metal ion and the CO₂-soluble chelating agent. The new entraining device maximizes contact between the ligand-rich CO₂ phase and the metal ion-rich aqueous phase. Iron(III) was extracted from water with the chelating agent 2,2,7-trimethyl-3,5-octanedione (H(tod)) and supercritical fluid CO₂ at 60 °C and 20.8 MPa. With entrainment, 79% of the iron was removed from the aqueous phase. This represents a three-fold increase in iron extraction efficiency over that of a static system.     

Determination of Microcystins in Cyanobacteria by Supercritical Fluid Extraction and Liquid Chromatography‐Tandem Mass Spectrometry

Abstract

A method for the detection of microcystins (microcystin LR, RR, and YR) in cyanobacteria by supercritical fluid extraction (SFE) and liquid chromatography‐mass spectrometry (LC/MS) has been developed. Supercritical fluids for the analytical extraction of nonvolatile, higher molecular weight compound, and microcystins from cyanobacteria were investigated. The microcystins included in this study are sparsely soluble in neat supercritical fluid CO₂. However, the microcystins was successfully extracted with a ternary mixture (90% CO₂, 9.5% methanol, 0.5% water) at 40°C and 250 atm. The polar carbon dioxide‐aqueous methanol fluid system gave high extraction efficiency for the extraction of the polar microcystins from cyanobacteria. The microcystins were determined by liquid chromatography‐tandem mass spectrometry (LC/MS/MS).     

Supercritical fluid solid chromatography using novel porous glassy carbon. Evaluation of retention mechanisms

The use of a porous glassy carbon (PGC) material as a packed-column SFC stationary phase has been previously demonstrated [1]. The material is further characterized in terms of its retention characteristics. The effects of variations in mobile phase composition, pressure, and temperature conditions are evaluated. Variation of temperature and pressure yielded expected results, specifically, decreased solute capacity factors with increased mobile phase density. The choice of supercritical fluid mobile phase allows the most notable control of solute retention; this was evaluated by adding low percentages of organic modifiers of varying molecular weights to the supercritical carbon dioxide mobile phase. PGC-SFC provides reversed phase characteristics similar to those found for PGC-HPLC. Porous glassy carbon has selectivity characteristics previously unavailable in supercritical fluid chromatography. Use of porous glassy carbon in supercritical fluid chromatography may provide distinct advantages in difficult analytical separations, allowing separations of molecules with only slight structural differences.     

Retention behavior of polycyclic aromatic hydrocarbons in supercritical fluid chromatography on a chemically bonded stationary phases based upon liquid-crystalline polymer

Summary The retention behavior of a set of polycyclic aromatic hydrocarbons in supercritical fluid chromatography have been studied on a chemically bonded stationary phase based upon a side chain liquid crystalline polymer (LCP) with carbon dioxide-based mobile phase. The effects of the mobile phase pressure, column temperature and amount of mobile phase organic modifier have been investigated in order to detect a possible structural change in the liquid crystal polymer linked to the silica support. The influence of these factors on the selectivity coefficients has also been studied. Two distinctive behaviors with temperature are noted at low pressure on the one hand and at higher pressure on the other. This change in behavior is based on the density of the supercritical CO₂ and the PAH volatility rather than on any specific stationary phase structural change. Both lower mobile phase pressure and amount of mobile phase modifier are required to obtain better selectivities. Better planarity recognition is observed in SFC than in HPLC with these new bonded liquid crystal stationary phases. The bonded liquid crystal phase is only weakly affected by the addition of organic modifier in the supercritical CO₂.     

Influence of temperature on FTIR spectra of various organic compounds analyzed by on-line carbon dioxide SFC/FTIR

The development of an on-line SFC-FTIR method with supercritical carbon dioxide as mobile phase requires information about the nature of the FTIR spectrum of a solute dissolved in supercritical or liquid CO₂. The wavenumber of maximum absorbance of the carbonyl stretching vibration was quantitatively studied versus temperature and CO₂ density. FTIR spectra recorded in supercritical or liquid CO₂ have been compared to their vapor-phase and condensed-phase spectra for various solutes.     

Addition of modifier in supercritical fluid chromatography using a microbore reciprocating pump

Summary Solubilities of polar analytes in supercritical CO₂ can be enhanced by the addition of polar modifiers. The addition of modifier into an SFC system using a low cost reciprocating pump has been studied. Two different mixing chambers were evaluated for mixing the supercritical CO₂ with modifier. It appeared that a mixing chamber with a packed bed was enough to reduce baseline noise from the modifier pump. Results from the effect of pressure and temperature with various modifier flow rates were obtained. High percentages of modifier (>15%) at a low CO₂ pressure (2000 psi) caused baseline instability. In addition, different I.D. columns were tested with the system and the effect of modifier compressiblity on detector noise was also studied. Several pharmaceutical compounds were separated to demonstrate system performance.     

Rapid Supercritical Fluid Chromatography Method for Separation of Chlorthalidone Enantiomers

Supercritical fluid chromatography employing chiral stationary phases is a popular separation technique to perform enantioselective separations. The main advantages of supercritical fluid chromatography are low analysis time, low consumption of organic modifiers, and therefore lower costs and higher environmental friendliness. A novel method for the separation of chlorthalidone enantiomers, widely used diuretic drug, is reported that clearly demonstrates the advantages of supercritical fluid chromatography. The effects of the amount and type of organic modifiers, temperature, and back pressure on enantioselectivity and resolution of the enantiomers were evaluated. The baseline separation was achieved in less than 2.5 min in the optimized system composed of Chiralpak AD column, mobile phase CO₂/MeOH 50/50 (v/v), temperature 40°C, a flow rate of 4.0 mL/min, and 120 bar back pressure. Moreover, enantiomers of chlorthalidone were determined in two commercially available pharmaceuticals. The proposed method may be easily transferred to a semi-preparative scale.     

A detection method for unified chromatography: Ion mobility monitoring

Ion mobility monitoring has been used for detection in gas, supercritical fluid, and liquid chromatography, illustrating its potential as a method of detection for unified chromatography. Applications presented include GC-IMD of dioxins in fly ash, SFC-IMD of vitamin E, and HPLC-IMD of alkylamines. Ion mobility spectra of several mixed supercritical fluid mobile phases are also presented. Use of methanol, acetonitrile, and dichloromethane as modifiers of supercritical carbon dioxide, and use of supercritical dichlorodifluoromethane and chlorodifluoromethane as mobile phases had little effect on the reactant ion pattern at the flow rates and concentrations used in this study. Only when acetone was used as a modifier of carbon dioxide did the positive reactant ions change significantly. No effect of modifiers or mobile phase was observed for the negative reactant ions.     

On-Line coupling of supercritical fluid extraction with high performance liquid chromatography

Supercritical fluid extraction was coupled directly with high performance liquid chromatograph. The system was evaluated for direct injection of supercritical CO₂ and modified supercritical CO₂ at high pressure and temperature onto a HPLC system with varying mobile phase compositions and flow rates. Injection of 9 μL supercritical CO₂ onto the HPLC using methanol/water mobile phases from 100% methanol to 80% with a flow of 1.0 mL/min did not adversely affect the baseline of UV detector. However at higher percentages of water, CO₂ solubility in the mobile phase decreased and caused baseline interferences on the UV detector. At higher HPLC mobile phase flow rates, supercritical CO₂ was injected to higher percentages of water without any effect on the UV baseline. Also, increasing the extraction pressure or modifier concentration did not change the results. Separations of polynuclear aromatic hydrocarbons and linear alkenebenzene sulfonate test mixtures were obtained using on-line SFE/HPLC interfaced system.     

Packed column supercritical fluid chromatography with light-scattering detection. II. Retention behaviour of squalane and glucose with mixed mobile phases

Summary Evaporative light scattering detectors can be used to detect organic substances without chromophoric groups in packed column supercritical fluid chromatography (SFC). A detector of this type has been used to detect squalane and glucose after SFC with various packed columns and binary mobile phases. In this study, the amount of organic modifier in carbon dioxide/modifier mixtures was varied. The results give further insight into the mechanisms that influence retention behaviour in packed column separations with super- and subcritical mobile phases. Squalane is an ideal non-polar test solute which shows long retention times on non-polar columns while its elution can be accelerated by non-polar modifiers in carbon dioxide. Glucose is an extremely polar solute containing hydroxyl groups. Elution of this sugar can be improved with polar modifiers. Column packings with polar end groups lead to high capacity ratios and long retention times for glucose. Most columns used in this study contained silica-based packing materials. For purposes of comparison, a polymeric packing (HEMA RP-18) was also employed.     

Enhanced fluidity liquid chromatography for hydrophilic interaction separation of nucleosides

The application of enhanced fluidity liquid (EFL) mobile phases to improving isocratic chromatographic separation of nucleosides in hydrophilic interaction liquid chromatography (HILIC) mode is described. The EFL mobile phase was created by adding carbon dioxide to a methanol/buffer solution. Previous work has shown that EFL mobile phases typically increase the efficiency and the speed of the separation. Herein, an increase in resolution with the addition of carbon dioxide is also observed. This increase in resolution was achieved through increased selectivity and retention with minimal change in separation efficiency. The addition of CO₂ to the mobile phase effectively decreases its polarity, thereby promoting retention in HILIC. Conventional organic solvents of similar nonpolar nature cannot be used to achieve similar results because they are not miscible with methanol and water. The separation of nucleosides with methanol/aqueous buffer/CO₂ mobile phases was also compared to that using acetonitrile/buffer mobile phases. A marked decrease in the necessary separation time was noted for methanol/aqueous buffer/CO₂ mobile phases compared to acetonitrile/buffer mobile phases. There was also an unusual reversal in the elution order of uridine and adenosine when CO₂ was included in the mobile phase.     

Structural features of binary mixtures of supercritical CO2 with polar entrainers by molecular dynamics simulation

Computer simulations of supercritical carbon dioxide and its mixtures with polar cosolvents: water, methanol, and ethanol (concentration, 0.125 mole fractions) at T = 318 K and ρ = 0.7 g/cm³ are performed. Atom-atom radial distribution functions are calculated by classical molecular dynamics, while the probability distributions of relative orientation of CO₂ molecules in the first and second coordination spheres describing the geometry of the nearest environment of CO₂ molecules and the trajectories of cosolvent molecules are found using Car-Parrinello molecular dynamics. Based on the latter, the conclusions regarding structure and interactions of polar entrainers in their mixtures with supercritical CO₂ are made. It is shown that the microstructure of carbon dioxide varies only slightly upon the introduction of cosolvents.     

Homogeneous and heterogeneous free radical polymerizations in environmentally responsible supercritical carbon dioxide

Fluoropolymers are used in many technologically demanding applications because of their balance of high-performance properties. A significant impediment to the synthesis of variants of commercially available amorphous fluoropolymers is their general insolubility in most solvents except chlorofluorocarbons (CFCs). The environmental concerns about CFCs can be circumvented by preparing these technologically important materials in supercritical fluids (SCFs). The homogeneous solution homo- and copolymerization of highly fluorinated acrylic, styrenic and olefinic monomers in supercritical carbon dioxide using free radical methods will be discussed [Science, 257 , 945 (1992)]. Detailed decomposition rates and efficiency factors will be presented for azobisisobutyronitrile (AIBN) in supercritical carbon dioxide and will be compared to conventional liquid solvents [Macromolecules, 26 , 2663 (1993)]. Additionally, viscosities of polymer solutions in supercritical CO₂ have been measured using a high pressure, falling cylinder viscometer. The results show that the polymer solution viscosities in supercritical CO₂ are an order of magnitude lower than with the same polymers in conventional organic solvents. The results from these homogeneous solution polymerization studies has allowed us to also consider heterogeneous polymerizations in a carbon dioxide continuous phase. Conventional emulsion polymerizations of unsaturated monomers are performed in either aqueous or organic dispersion media with addition of surface active agents (surfactants) to stabilize the colloidal dispersion that forms. With free radical initiators that are preferentially soluble in the continuous phase, high rates of polymerization and high molar mass polymers can be obtained simultaneously. Herein we describe an environmentally responsible alternative to aqueous and organic dispersing media for emulsion polymerizations which utilizes supercritical carbon dioxide, in conjunction with molecularly engineered free radical initiators and amphiphilic molecules that are specifically designed to be interfacially active in CO₂. Conventional lipophilic monomers, exemplified by methyl methacrylate and styrene, can be polymerized heterogeneously using a fluorinated azo-initiator in supercritical CO₂ in the presence of added surfactant to form stable emulsions that result in submicron size particles. Detailed surfactant and initiator syntheses and phase behavior will also be discussed.     

Fluorinated surfactants in supercritical CO2

Liquid or supercritical carbon dioxide has important environmental and economic advantages over petrochemical solvents currently used for industrial processes. However, low solubility in CO₂, particularly of polar compounds, is a hurdle to its implementation as an acceptable alternative. These solubility problems have been overcome by employing specialised fluorinated surfactants to stabilise water nano-droplets as water-in-supercritical/liquid CO₂ microemulsions. Such novel microemulsions can now facilitate innovative ‘green-and-clean’ applications of carbon dioxide technology.     

Overview of the retention in subcritical fluid chromatography with varied polarity stationary phases

Retention and separation of achiral compounds in supercritical fluid chromatography (SFC) depend on numerous parameters: some of these parameters are identical to those encountered in HPLC, such as the mobile phase polarity, while others are specific to SFC, as the density changes of the fluid, due to temperature and/or pressure variations. Additional effects are also related to the fluid compressibility, leading to unusual retention changes in SFC, for instance when flow rate or column length is varied. These additional effects can be minimised by working at lower temperatures in the subcritical domain, simplifying the understanding of retention behaviours. In these subcritical conditions, varied modifiers can be mixed to carbon dioxide, from hexane to methanol, allowing tuning the mobile phase polarity. With nonpolar modifiers, polar stationary phases are classically used. These chromatographic conditions are close to the ones of normal-phase LC. The addition of polar modifiers such as methanol or ACN increases the mobile phase polarity, allowing working with less polar stationary phases. In this case, despite the absence of water, retention behaviours generally follow the rules of RP LC. Moreover, because identical mobile phases can be used with all stationary phase types, from polar silica to nonpolar C18-bonded silica, the classical domains, RP and normal-phase, are easily brought together in SFC. A unified classification method based on the solvation parameter model is proposed to compare the stationary phase properties used with the same subcritical mobile phase.     

The phase behavior of fluorinated diols,divinyl adipate and a fluorinated polyester in supercritical carbon dioxide

The use of supercritical carbon dioxide as a reaction medium for polyester synthesis is hindered by the low solubility of diols in CO₂. However, it has been previously demonstrated that fluorinated compounds can exhibit greater miscibility with carbon dioxide than their hydrocarbon analogs. Therefore, the phase behavior of fluorinated diols and divinyl adipate (DVA), an activated diester, in supercritical carbon dioxide has been investigated at 323 K. The phase behavior of equimolar mixtures of DVA with the most carbon dioxide-soluble diol, 3,3,4,4,5,5,6,6-octafluorooctan-1,8-diol (OFOD), was also determined. The solubility of a polyester synthesized from DVA and 2,2,3,3-tetrafluoro-1,4-butanediol (TFBD) was found to be less CO₂-soluble than its monomers. DVA was much more soluble in CO₂ than any of the fluorinated diols, therefore, no attempt was made to fluorinate the DVA structure. Because both substrates and polyester product were soluble in carbon dioxide, the enzymatic synthesis of a fluorinated polyester from DVA and octafluorooctandiol was performed in supercritical carbon dioxide, resulting in a polymer with a weight average molecular weight of 8232 Da.     

Palladium‐catalyzed carbonylation of primary amines in supercritical carbon dioxide

The chemoselectivity of the palladium‐catalyzed carbonylation of amines was affected by the addition of MeOH in supercritical carbon dioxide. The results show different selectivity in supercritical carbon dioxide CO₂(sc) from that in alcohol. Methyl carbamate and its derivatives were obtained in high yields in CO₂(sc).     

Clinical and pharmaceutical applications of packed-column supercritical fluid chromatography

Packed-column supercritical fluid chromatography (pSFC) is a fast separation technique that combines the properties of HPLC and GC. pSFC with carbon dioxide as the mobile phase and packed silica column as the stationary phase possesses the properties of normal phase mechanism; however, the addition of modifiers to the mobile phase allows the separation of relatively polar compounds. In spite of its many positive attributes, pSFC has not been widely used in areas such as proteomics, where methods such as HPLC dominate. Packed column SFC has been extensively used in clinical and pharmaceutical laboratories, especially for separation of nonpolar and chiral drugs. This review will discuss recently published applications of pSFC, with a specific focus on its advantages and limitations for the analysis of pharmaceuticals with varying chemical properties.