A supercritical fluid workflow for the quality assessment of herbal drugs and commercial preparations from Rhodiola rosea.

Comparison of ultra-high performance supercritical fluid chromatography and ultra-high performance liquid chromatography for the analysis of pharmaceutical compounds

The importance of hops (the flowers of Humulus lupulus) as food and an herbal remedy is reflected by a large number of analytical methods published. However, supercritical fluid chromatography, a highly efficient, rapid, and "green" separation technique, has not been considered for hops samples so far. This prompted us to establish the first supercritical fluid chromatography-based protocol for the separation, identification, and quantitation of five prenylated constituents of hops. Hulupinic acid ( 1: ), a prominent oxidation product of hop acids, three flavanones, i.e., 8-prenylnaringenin ( 2: ), 6-prenylnaringenin ( 3: ), and isoxanthohumol ( 4: ), as well as the chalcone xanthohumol ( 5: ) could be baseline separated in lessthan 5 minutes using a Viridis BEH 2-EP column (3.0 × 100 mm; 1.7 µm particle size) and a mobile phase consisting of CO2 and isopropanol. Good results regarding selectivity, accuracy (recovery rates: 85.0 - 113.1%), precision (intra-day ≤ 2.1%, inter-day ≤ 3.5%), and linearity (R2 ≥ 0.99) were obtained for both photodiode array and mass detection. The lowest detection limit at 220 nm was at 0.1 µg/mL ( 1, 3: , and 4: ), with mass detection even at 0.001 µg/mL ( 4: ). As an application example of the validated method, the five hops constituents were quantified in three dietary supplements, one herbal medicinal product, and two batches of hop flowers (Lupuli flos). In most samples analyzed, the major component was 5: (0.01 - 1.02%), whereas the major component in Lupuli flos samples was compound 1: (0.12 - 0.21%). This protocol offers a fast and environmentally friendly alternative to liquid chromatography for the quality control of hops.

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.

Development and validation of ultra-high performance supercritical fluid chromatography method for determination of illegal dyes and comparison to ultra-high performance liquid chromatography method

Phenolic acid as an important active ingredient of traditional Chinese medicine, have received extensive attention due to its various biological activities and pharmacological effects. However, the presence of isomers makes the separation of phenolic acids challenging. In the present study an environmentally friendly, sustainable, lower cost, and green that reduce amounts of organic solventsultra-performance convergence chromatography (UPC2) method was developed for separation and quantification eight phenolic acids. Ultra-high performance supercritical fluid chromatography combines the advantages of supercritical fluid chromatography and ultra high performance liquid chromatography technologies with shorter analysis time, higher resolution and better environmental compatibility. The effects of different chromatographic column, co-solvent, column temperature and back pressure were studied. The BEH column was provided the most suitable separation withgradient elution in 20 min using CO2 as the mobile phase and methanol/acetonitrile (70/30, v/v) with 2% TFA as the co-solvent at a flow rate of 0.4 mL/min. A better and more symmetrical peak shape were obtained. Subsequently, the established method was applied to the quantification of phenolic acids in raw and processed Tussilago farfara L. and validated according to FDA guidelines, showed good precision, accuracy, repeatability and linearity in the concentration range. Our findings confirm that the method could be used as an efficient method for exploring phenolic acids of the quality and separation of traditional Chinese medicine.

Comparison of ultra-high performance supercritical fluid chromatography and ultra-high performance liquid chromatography for the separation of spirostanol saponins

Picea abies (L.) H. Karst. (Pinaceae) is native to Northern, Central and Eastern Europe. The fast-growing tree reaches up to 50 m in height, has modest nutritional requirements and depends on sufficient water supply. The conifer, commonly called Norway spruce, produces exudates which are traditionally used to treat skin wounds in Northern European countries. Major bioactive constituents of the conifer oleoresin are diterpene resin acids (DRAs) of the abietane and the pimarane type. To assure consistent pharmaceutical quality of Norway spruce balm and commercial products thereof, an analytical method for the quantitation of DRAs is the prerequisite. However, high structural similarity among DRAs and their poor UV absorption makes chromatographic separation and detection challenging: Conventional liquid chromatography systems often fail to achieve sufficient separation, moreover, they are not sustainable. Gas chromatography on the other hand requires time-consuming derivatization prior to unacceptably long analyses (>60 min). These drawbacks prompted the development of the first validated supercritical fluid-based protocol for the separation and quantitation of eight DRAs, i.e., pimaric acid (1), sandaracopimaric acid (2), palustric acid (3), isopimaric acid (4), levopimaric acid (5), abietic acid (6), dehydroabietic acid (7), and neoabietic acid (8). By using an ultra high-performance supercritical fluid chromatography (UHPSFC) device hyphenated to a quadrupole mass detector, the DRAs were separated in less than 20 min on a Torus 2-Picolylamin (2-PIC) column (3.0 mm × 100 mm; 1.7 µm particle size) applying supercritical CO2 and ethanol as mobile phase. Regarding selectivity, accuracy (recovery rates: 87–108%), intermediate precision (between 6.6 and 11.1%), and linearity (R2 ≥ 0.99; linear between 0.75 μg/ml and 2.5 mg/ml), results were obtained in line with ICH guidelines. The lowest limit of detection (LOD) was at 0.75 μg/ml (7) and the lowest limit of quantitation (LOQ) at 2 μg/ml (8). As application examples, 22 Norway spruce balm samples and five commercial products were analyzed. The here presented protocol not only simplifies and shortens the analytical workflow, but also reduces the amount of organic solvent waste by about two thirds compared to conventional liquid chromatographic set-ups. These advantages qualify this fast and efficient method as an ideal tool for an environmentally friendly quality control of traditionally used wound-healing Norway spruce balm products.

Supercritical fluid chromatography (SFC), which employs pressurized carbon dioxide as the major component of the mobile phase, has been known for several decades but has faced a significant resurgence of interest in the recent years, thanks to the development of modern instruments to comply with current expectations in terms of robustness and sensitivity. This review is focused on the recent literature, specifically since the introduction of modern systems but in relation to older literature, to identify the changing trends in application domains. Typically, natural products, bioanalysis, food science, and environmental analyses are all strongly increasing. Together with reduced extra-column volumes in the instruments, the advent of sub-2-μm particles and superficially porous particles in the stationary phases is favoring ultra-high-performance SFC (UHPSFC) allowing for improved resolution and faster analyses, but without the constraints of viscous liquids encountered in ultra-high-performance liquid chromatography (UHPLC). Hyphenation to mass spectrometry is also more frequent and opened the way to new application domains, and raises different issues from liquid chromatography mobile phases, especially due to decompression of carbon dioxide. It is also shown that the frontiers between SFC and HPLC are fading, as switching from one method to the other, even within the course of a single analysis, is facilitated my modern instruments. The present review is not intended to be exhaustive but rather giving a snapshot of recent trends in supercritical fluid chromatography, based on the observation of about 500 papers published in English-written peer-reviewed journals from 2014 to 2018. Graphical abstract ᅟ.

Boswellic acids, a class of triterpenes, are the bioactive constituents in Indian frankincense, an herbal drug with pronounced anti-inflammatory activity. In this study their separation and quantification in B. serrata extracts is reported for the first time by using Supercritical Fluid Chromatography. Under optimized conditions, i.e. a Viridis HSS C18 SB column and carbon dioxide, methanol, acetonitrile and ammonium hydroxide as mobile phase, six boswellic acids could be separated in under 6 min. The assay fulfilled all validation criteria with coefficients of determination higher than 0.999, a wide linear range (30–1000 μg/mL), recovery rates from 97.1–103.0 %, excellent precision, and detection limits typical for SFC with UV-detection (≤ 5.5 μg/mL). The method could easily be hyphenated to mass spectrometry, which was helpful to tentatively assign further compounds (mainly derivatives of tirucallic acid) and to increase the assay’s sensitivity. Its practical applicability was confirmed by analyzing several commercial products, which mainly contained β-boswellic acid as dominant triterpene, yet in extremely variable amounts ranging from 0.9 to 16.9 %.

Enantiomeric separation and quantification of R/S-amphetamine in urine by ultra-high performance supercritical fluid chromatography tandem mass spectrometry

A rapid and simple ultra-high-performance supercritical fluid chromatography (UHPSFC) with photodiode array (PDA) method was developed and validated for simultaneous determination of eight Sudan dyes in chili oil. In particular, a pair of isomer, Sudan red B and Sudan IV, was included in the analysis. After being diluted with dichloromethane, the analytes were separated on an Acquity UPC2 HSS C18 SB column with gradient elution using CO2 as the mobile phase and acetonitrile/methanol (v/v, 45/55, containing 0.1% formic acid) as the organic modifier. Analytes were quantified by external calibration curves over ranges of 0.5–50 mg/L, with correlation coefficients above 0.999. The method gave recoveries of the target compounds (spiked at levels of 1, 5, and 25 mg/kg) ranging from 82.6 to 108.3%, with intraday and interday relative standard deviations of less than 8.0% and 8.6%, respectively. The limits of detection (LODs) and the limits of quantification (LOQs) for eight dyes were from 0.10 to 0.30 mg/kg and 0.30–1.00 mg/kg, respectively. This method was applied for the analysis of chili oil samples collected from the supermarket in Beijing. This validated that the UHPSFC-PDA method provides a useful strategy for the simultaneous determination of Sudan dyes in chili oil for routine analysis.

A simple, reliable method for the detection of free and modified Fusarium mycotoxins in beer using state-of-the-art ultra-high-performance supercritical fluid chromatography (UHPSFC) with low-resolution tandem MS (MS/MS) is presented in this paper. The UHPSFC-MS/MS method was developed for nivalenol, deoxynivalenol, 15-acetyl-deoxynivalenol, 3-acetyl-deoxynivalenol, deoxynivalenol-3-glucoside, HT-2 toxin, T-2 toxin, T-2 toxin-3-glucoside, neosolaniol, diacetoxyscirpenol, zearalenone, α-zearalenol, and β-zearalenol and their internal standards deepoxy-deoxynivalenol and zearalanone. Due to the broad range of the physicochemical properties of the aforementioned, the sample preparation step was minimized to avoid analyte losses. Extraction with acetonitrile-water-acetic acid (79 + 20 + 1, v/v/v) and hexane in combination with solid-phase extraction (C18) was followed by a filtration step. After filtration, the extract was evaporated, and the remaining residue was redissolved in a mobile phase for injection (methanol-water; 90 + 10, v/v). A mobile phase consisting of supercritical CO2 and a small portion of methanol was used. The developed multimycotoxin method permits the simultaneous determination of multiple fusariotoxins in an one-step chromatographic run using UHPSFC-MS/MS. SFC is a promising strategy; however, the retention mechanism is complex, leading to the unpredictable nature of elution and to some mycotoxins not being retained on the column. This restricts the applicability of UHPSFC in multimycotoxin analyses. The present study is the first report on the use of UHPSFC for the analysis of free and modified Fusarium mycotoxins.

Fast analysis of derivatives of polycyclic aromatic hydrocarbons in soil by ultra-high performance supercritical fluid chromatography after supercritical fluid extraction enrichment

One of the most significant techniques for column chromatography, supercritical fluid chromatography (SFC), combines the principles of gas chromatography (GC) and high-performance liquid chromatography (HPLC). As the mobile phase, a supercritical fluid is utilised. The mobile phase is mostly CO2. Supercritical fluids combine advantageous characteristics of the liquid and gas phases. The equipment required for Supercritical Fluid Chromatography is versatile as it is compatible with multiple detectors. Mainly Flame Ionization Detector is used in Supercritical Fluid Chromatography. Recently, a number of researchers have demonstrated that the use of alternative solvents or the addition of modifiers to SC-CO2 improves the performance of supercritical fluid technology. UHPSFC-MS (Ultra-High Performance Supercritical Fluid Chromatography–Mass Spectrometry)is one of the recent advances in Supercritical Fluid Chromatography. For separations involving non-volatile or thermolabile species that cannot be separated by GC or LC, supercritical fluid chromatography is frequently utilised. SFC has been applied to a wide range of materials including natural products, pharmaceuticals, foods, pesticides, herbicides, surfactants, polymers and polymer additives, fossil fuels, petroleum, explosives, and propellants.

Comparison of liquid chromatography and supercritical fluid chromatography coupled to compact single quadrupole mass spectrometer for targeted in vitro metabolism assay