Determination of alcohol polyether average molar oligomer value/distribution via supercritical fluid chromatography coupled with UV and MS detection

Supercritical fluid chromatography (SFC) was used for the analysis of ethoxylated and propoxylated surfactants. Samples were derivatized to phenylated silyl ethers with a disilazane-chlorosilane mixture. Addition of a phenyl group to the surfactant allowed UV-absorbance detection of each oligomer. Acetonitrile and methanol were evaluated as mobile phase modifiers. Better peak shape was realized with methanol-modified CO2 on an octadecyl silica bonded phase than with acetonitrile-modified CO2. Peak assignments were made via SFC coupled with electrospray ionization-mass spectrometry (ESI-MS) in the positive ion mode. A sulfonamide-embedded alkyl stationary phase was also evaluated for separation of the derivatized samples. SFC-UV and SFC-ESI-MS data were jointly used for calculation of average molar oligomer values which were then compared to values calculated from 1H NMR data of non-derivatized samples. The derivatization or separation method using the sulfonamide embedded phase required no preliminary cleanup and yielded reproducible oligomer values that were consistent with those of the manufacturer's nominal values.     

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.     

Construction of a new interface for comprehensive supercritical fluid chromatography x reversed phase liquid chromatography (SFC x RPLC)

The design of a new interface to hyphen high efficiency supercritical fluid chromatography (SFC) and fast RPLC in a comprehensive configuration is described. SFC x RPLC is a viable alternative to normal phase (NP) LC x RPLC and is characterized by high orthogonality. Compared to NPLC x RPLC an additional advantage is the expansion of supercritical carbon dioxide (CO(2)) when exposed to atmospheric pressure leading to fractions consisting of solvents that are miscible with the second dimension RPLC mobile phase. The interface consists of a two-position/ten-port switching valve equipped with two packed octadecyl silica (C(18)) loops for effective trapping and focusing of the analytes after elution from the SFC dimension. The addition of a water make-up flow to the SFC effluent prior to entering the loops is of fundamental importance to efficiently focus the solutes on the C(18) material and to reduce interferences of expanded CO(2) gas on the second dimension separation. The features of the system are illustrated with the analysis of a lemon oil sample.     

Development of an orthogonal method for mometasone furoate impurity analysis using supercritical fluid chromatography

While supercritical fluid chromatography (SFC) has received great popularity in chiral separation and purification, it has rarely been used for trace level pharmaceutical impurity analysis, partially due to the limitation of instrument sensitivity. In this study, a packed column SFC method has been developed for the quantitative analysis of mometasone furoate and its trace level impurities. The UV detection was optimized to improve the sensitivity by 2-4 fold. In combination with an increased sample concentration, this SFC method is capable of trace level (0.05% of the active) analysis of the impurities. The SFC method used a silica column and a mobile phase consisting of CO(2) and methanol. The new method provides an orthogonal selectivity complementary to the reversed phase HPLC (RP-HPLC) method. All of the impurities and the active were baseline separated within 12 min on SFC, which is less than one third of the RP-HPLC method run time. The method was also partially validated for linearity, accuracy, precision (repeatability), and limit of quantitation. This study demonstrated that the SFC method, with improved sensitivity, can be a valuable tool to provide orthogonal selectivity for trace level impurity separation. With further validation, the method may be suitable for release testing and stability testing for mometasone furoate drug substance.     

Retention mechanisms in super/subcritical fluid chromatography on packed columns

Whereas the retention rules of achiral compounds are well defined in high-performance liquid chromatography, on the basis of the nature of the stationary phase, some difficulties appear in super/subcritical fluid chromatography on packed columns. This is mainly due to the supposed effect of volatility on retention behaviours in supercritical fluid chromatography (SFC) and to the nature of carbon dioxide, which is not polar, thus SFC is classified as a normal-phase separation technique. Moreover, additional effects are not well known and described. They are mainly related to density changes of the mobile phase or to adsorption of fluid on the stationary phase causing a modification of its surface. It is admitted that pressure or temperature modifications induce variation in the eluotropic strength of the mobile phase, but effects of flow rate or column length on retention factor changes are more surprising. Nevertheless, the retention behaviour in SFC first depends on the stationary phase nature. Working with polar stationary phases induces normal-phase retention behaviour, whereas using non-polar bonded phases induces reversed-phase retention behaviour. These rules are verified for most carbon dioxide-based mobile phases in common use (CO(2)/MeOH, CO(2)/acetonitrile or CO(2)/EtOH). Moreover, the absence of water in the mobile phase favours the interactions between the compounds and the stationary phase, compared to what occurs in hydro-organic liquids. Other stationary phases such as aromatic phases and polymers display intermediate behaviours. In this paper, all these behaviours are discussed, mainly by using log k-log k plots, which allow a simple comparison of stationary phase properties. Some examples are presented to illustrate these retention properties.     

Supercritical fluid chromatographic solvatochromic modifier polarity studies

To understand the chromatographic process as a whole, whether it be for gas chromatography (GC), liquid chromatography (LC), or supercritical fluid chromatography (SFC), one needs to know the chemical and physical nature of the mobile and stationary phases and also the interactions that take place between analytes (solutes) and the two phases. An approach towards Investigating the ways that stationary and mobile phases contribute to chromatographic retention Involves exploring the effects of solvent polarity on the strength of the mobile phase. In SFC this could involve determining the polarity of several different modifier/carbon dioxide mobile phases. In this paper, the use of a solvatochromic indicator to learn more about the effects of SFC modifier/mobile phase polarity will be investigated and discussed using several different modifiers and a diolmodified silica column.     

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 geometrical isomers of sesquiterpene and diterpene alcohols on bare silica by carbon dioxide supercritical fluid chromatography

Summary The application of supercritical fluid chromatography (SFC) with modified carbon dioxide to the separation of geometrical isomers of several sesquiterpene or diterpene alcohols is reported. The chromatographic system consists of bare silica (250×4.6 mm i. d.) with carbon dioxide-methanol mobile phase and UV detection. A mixture of sesquiterpene alcohols, (2E-6E)-farnesol, -bisabolol, (E)-nerolidol and (Z)-nerolidol, and also of two diterpene alcohols, (R)- and (S)-geranyllinalol, was separated by SFC using a mixture of carbon dioxide modified with 0.5–1.0% methanol as mobile phase.     

A simultaneous packed column supercritical fluid chromatographic method for ibuprofen, chlorzoxazone and acetaminophen in bulk and dosage forms

A reproducible and efficient method for the separation and estimation of ibuprofen, chlorzoxazone and acetaminophen has been developed using packed column supercritical fluid chromatography (SFC). The separations were performed on an ODS-RP JASCO column employing methanol modified supercritical fluid CO(2) as the mobile phase. The densities and polarities of the mobile phase were optimised from the effects of pressure, temperature and modifier concentration on retention times. In addition a flow programming of the mobile phase helped to obtain better resolution and a faster elution for acetaminophen. The analytes were detected using a uv detector at 254 nm. The study includes a successful attempt at quantitation of the 3 drugs. Chromatographic figures of merit, linear dynamic range, limit of quantitation (LOQ), precision and accuracy etc. were determined to assess the viability of the method. The method has been extended to commercial dosage forms containing all 3 drugs.     

高效液相色谱中的手性两相协同作用

在高效液相色谱中,以纤维素三苯基甲酸酯、纤维素三苯基氨基甲酸酯为手性固定相,以正己烷-异丙醇(体积比为9∶1)为流动相,柱温25 ℃,分别以β-环糊精（β-CD）、2,6-二甲基-β-环糊精(DM-β-CD)、2,3,6-三甲基-β-环糊精(TM-β-CD)为流动相添加剂,分离了DL-色氨酸和DL-苯丙氨酸两种手性化合物,考察是否存在手性固定相和手性添加剂的协同作用。实验结果表明,在流动相中添加β-CD或DM-β-CD至饱和,协同作用不明显;在流动相中添加少量TM-β-CD(即浓度小于0.60 mmol/     

Separation of Natural Food Pigments in Saponified and Un-Saponified Paprika Oleoresin by Ultra High Performance Supercritical Fluid Chromatography (UHPSFC)

The natural pigments in paprika were rapidly and efficiently separated by ultra high performance supercritical fluid chromatography. The separation of both un-saponified and saponified mixtures of paprika oleoresin were optimized, with run times of 10.6 min. Three different C18 columns, a cyano, silica and diol column, all 3 × 100 mm, with 1.8 μm particles were compared. The best separation for the un-saponified sample was found with an SB-C18 column, while the saponified samples were best separated on a bare silica, RX-Sil column. A SB-CN column allowed near optimum separation of both the unsaponified, and saponified samples, with similar run times. The best mobile phase was carbon dioxide (CO₂) modified with isopropyl alcohol (IPA), with a composition gradient. Fingerprints of several commercial pepper products indicated that one appeared to be colored with artificial dyes, while the color of a chili powder may have been enhanced with a paprika extract. Spectra, using CO₂ with IPA as modifier, produced a single maximum at 453 nm, which appears to represent up to a 30 nm solvatochromic shift from the maxima in most organic solvents. Acetonitrile (ACN) as modifier produced spectra with two maxima and a similar solvatochromic shift. These results appear to be the first on saponified paprika oleoresin samples using SFC. It is also the first detailed report on the separation of un-saponified samples. The results are up to six times faster than comparable results by HPLC. It appears that SFC is a viable, superior alternative to HPLC for the analysis of this important commercial product, without using ACN, or chlorinated solvents.     

饲料中D-泛酸钙的超临界流体色谱测定

采用超临界流体色谱(SFC)测定了饲料中D-泛酸钙的含量.在二氧化碳流动相中添加14.85%甲醇改性剂及0.15%三氟乙酸添加剂,流速2 mL/min,在Kromasil ODS柱上分离,检测波长为210 nm.结果表明D-泛酸钙能在3 min内完成分离,在1.0 ～25.0 mg/L范围内,样品的质量浓度与色谱峰面积呈良好的线性关系(r=0.999 5).方法的精密度良好,相对标准偏差在2.3%以内,回收率为89% ～98%.方法操作简单,结果准确,适合普及应用.     

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.     

Enantioseparation of 1-phenyl-1-propanol on Chiralcel OD by supercritical fluid chromatography. I. Linear isotherm

The supercritical fluid chromatography (SFC) separation of the enantiomers of 1-phenyl-1-propanol on the chiral stationary phase Chiralcel OD under linear conditions is studied. Supercritical CO2 modified with methanol is used as a mobile phase. The effect of modifier concentration, pressure and temperature is studied. An empirical isotherm to account for the effect of density of the mobile phase and modifier concentration has been used to model the experimental results. It was observed that the selectivity and resolution were higher at 30 degrees C as compared to those at 40 degrees C.     

Direct resolution of racemic compounds on chiral microbore columns by sub- and supercritical fluid chromatography

Fast resolutions of racemic compounds (sulfoxides, amino alcohols, and α-methylarylacetic acids derivatives) were achieved on a chiral microbore column using carbon dioxide and a polar methanol/dioxane modifier. The stationary phase used in this study contains the 3,5-dinitrobenzoyl derivative of R,R(?)-1,2-diaminocyclohexane (DACH-DNB) as the chiral moiety, anchored to a silica gel surface by covalent bonds. Both thermodynamic and kinetic separation performances were improved by using a super- or subcritical carbon dioxide mobile phase (SFC, SubFC).     

Comparison of titania, zirconia, and silica stationary phases for separating diesel fuels according to hydrocarbon group-type by supercritical fluid chromatography

Silica, zirconia, and titania columns were compared for their ability to separate diesel samples into saturates, mono-, di-, tri-, and polyaromatics by supercritical fluid chromatography (SFC) using pure CO(2) according to ASTM method D 5186-03. A titania column coupled in series to a silica column was found to provide the highest overall group-type resolutions based on 20 model compounds, with resolutions as high as 14.7 for saturates versus monoaromatics and 11.9 for monoaromatics versus diaromatics. An oilsands-derived Synfuel light diesel, a commercial Ontario diesel, and a heavy Shell Canada Ltd. diesel blending feedstock were studied on a titania-silica coupled column as well as on a conventional silica column. The Synfuel results were similar (within 0.5 mass%) on both the conventional bare silica column and the titania-silica coupled column. The heavier commercial diesel and diesel blending feedstock samples yielded different results on the titania-silica coupled column compared to the silica column alone, demonstrating the importance of achieving the highest possible resolutions when baseline separation of group-types is not obtained.     

基于巯基-烯点击化学法的β-环糊精固定相多模式色谱保留行为研究

基于巯基硅胶与单取代-6A-烯丙氨基-β-环糊精的巯基-烯点击化学反应,制备了β-环糊精(Click TE-CD)共价键合固定相。元素分析结果表明β-环糊精被成功键合到硅胶表面。以黄酮苷类化合物为模型,考察了Click TE-CD固定相在亲水、反相和超临界流体色谱等分离模式下的色谱保留行为。黄酮苷类化合物保留时间随流动相中乙腈含量的变化呈现典型的U型曲线,表明Click TE-CD固定相具有亲水/反相的双重保留特性。应用几何学方法测得Click TE-CD固定相在反相/亲水、亲水/超临界、反相/超临界混合模式下的正交性分别为69.8%、50.8%、50.8%。对比复杂中药样品降香提取物在反相、亲水、超临界等模式下的分离情况,结果表明Click TE-CD固定相在分离中药复杂样品方面具有极大潜力,可以在一根色谱柱上通过分离模式的改变,实现二维液相色谱的分离。Click TE-CD固定相不同分离模式的分离性能和较好的正交性表明该固定相具有在液相色谱方法发展和二维液相色谱分离方面应用的潜力。     

Kinetic behaviour in supercritical fluid chromatography with modified mobile phase for 5 μm particle size and varied flow rates

After much development of stationary phase chemistry, in recent years the focus of many studies in HPLC has shifted to increase the efficiency and analysis speed. Ultra high pressure liquid chromatography (UHPLC) using sub-2 μm particles, and high temperature liquid chromatography (HTLC), using temperatures above 100°C have received much attention. These new approaches allow the use of flow rates higher than those classically used in HPLC, reducing the analysis duration. Due to the low viscosity of supercritical fluids, high velocities, i.e. high flow rates, can be achieved with classical pumping systems typically used in supercritical fluid chromatography (SFC). The effects of the flow rate increase with CO(2)/methanol mobile phase was studied on the inlet pressure, t(0), the retention factor of the compounds, and on the efficiency. Simple comparisons of efficiencies obtained at varied temperature between SFC and HPLC, with a packed column containing 5 μm particles, show the greater kinetic performances achieved with the CO(2)/methanol fluid, and underline specific behaviours of SFC, occurring for high flow rates and sub-ambient temperature. Some values (N/t(0)) are also compared to UHPLC data, showing that good performance can be achieved in SFC without applying drastic analytical conditions. Finally, simple kinetic plots (t(0) vs N) at constant column length are used to select combinations of temperature and flow rate necessary to achieve a required theoretical plate number.     

A crosslinked chiral stationary phase for the separation of enantiomers by gas and supercritical fluid chromatography

Crosslinking experiments for the chiral stationary phase OV-225-L-Val-t-butylamide within both fused silica and glass capillary columns have been carried out. Amino acid enantiomers were separated on crosslinked columns by both GC and SFC methods. In SFC, the a values of amino acid enantiomers are independent of the density of the mobile phase, and they are hig her than those obtained by GC for the tested enantiomers with the same column due to the lower column temperature used in SFC.     

Investigation of antioxidants and UV stabilizers from plastics. Part I: Comparison of HPLC and SFC; preliminary SFC/MS study

Synthetic mixtures of additives (antioxidants, UV stabilizers) used in the manufacture of food packaging materials were separated by normal phase HPLC under isocratic or gradient elution, and by capillary SFC using CO₂ as mobile phase. SFC proved to be more efficient for the separation and determination of additives with a wide range of boiling points and for substances lacking a UV chromophoric moiety. Combined SFC/MS, using a direct coupling interface, produced clear charge transfer spectra resembling EI spectra and proton exchange CI spectra when ammonia was added to the ion source.