Impact of injection solvents on supercritical fluid chromatography.

Even though there has been a rapid development in instrumentation and applications of supercritical fluid chromatography (SFC), relatively little is known about retention mechanisms compared to high-performance liquid chromatography (HPLC). Much effort has been made to characterize the influence of injection solvents on chromatographic efficiency in HPLC, however has been left rather uninvestigated in the domain of SFC. In this study properties of different injection solvents have been studied and correlated with properties of seven various analytes on three different columns, a C18, a 2-ethylpyridine and a bare-silica column. Aided by calculations of correlation coefficients and principal component analysis (PCA), the physical properties of injection solvents and the interactions between injection solvent, solute and stationary phase were investigated. The findings of this work shows that interactions capable of masking accessible silanol groups on a C18 column are of importance in order to maximize the plate number. While solvents with dipolar and hydrogen bond interaction properties are associated negatively with chromatographic efficiency using polar columns. Properties such as molar density, vapor pressure and boiling point were related to sharper peaks, mostly likely because of solubility issues of the injection solvent into the methanol-modified carbon dioxide. However, no additional solubility due to hydrogen interactions between the injection solvent and the carbon dioxide in SFC was observed. Surface tension and viscosity was not particularly associated with a decrease in plate numbers. By increasing the injection volume a stronger correlation between solubility related properties and plate numbers were obtained. Additional experiments showed that the resistance in solubility became an issue when performing partial-loop injection where additional washing solvent entered the system, thus providing broadened peaks

Pressurised hot water extraction in continuous flow mode for thermolabile compounds: extraction of polyphenols in red onions

Abstract Extraction and analysis of labile compounds in complex sample matrices, such as plants, is often a big analytical challenge. In this work, the use of a "green and clean" pressurised hot water extraction (PHWE) approach performed in continuous flow mode is explored. Experimental data for extraction and degradation kinetics of selected compounds were utilised to develop a continuous flow extraction (CFE) method targeting thermolabile polyphenols in red onions, with detection by high-performance liquid chromatography (HPLC)-diode array detection (DAD)-mass spectrometry (MS). Water containing ethanol and formic acid was used as extraction solvent. Method performance was focused on extraction yield with minimal analyte degradation. By adjusting the flow rate of the extraction solvent, degradation effects were minimised, and complete extraction could be achieved within 60 min. The CFE extraction yields of the polyphenols investigated were 80-90 % of the theoretically calculated quantitative yields and were significantly higher than the yields obtained by conventional methanol extraction and static batch extraction (70-79 and 58-67 % of the theoretical yields, respectively). The precision of the developed method was lower than 8 % expressed as relative standard deviation

Surface Sampling and Automated Liquid Membrane Extraction of Fungicides

A number of approaches for fungicide determination are presented. A surface sampling technique is utilised for the determination of fungicide foliar residues on greenhouse surfaces, mainly leaves. The procedure is based on solvation of the fungicides on the leaf surface with ethanol and HPLC analysis. It was applied to studies of the environment in greenhouses including investigations of decline rates of the fungicides. Occupational hazards, such as respiratory exposure and dermal uptake were investigated. Another environmental aspect on application of fungicides is the discharge of the compounds and their degradation products to water recipients. The actual concentrations are often quite low (usually below 1 µg/L) necessitating sensitive and selective analytical procedures involving enrichment. This can be met by membrane-based extraction techniques also providing possibilities for automation and closed systems. The supported liquid membrane (SLM) extraction and microporous membrane liquid-liquid extraction (MMLLE), are used for sample preparation of natural water samples. Automated systems on-line to reverse- and to normal phase HPLC have been realised. A combination of these techniques offers extraction conditions, which makes it possible to determine a wide variety of compounds

Determination of bile acids by hollow fibre liquid-phase microextraction coupled with gas chromatography.

A method based on hollow-fibre liquid phase microextraction combined with gas chromatography was developed for determination of specific bile acids in caecal materials of rats. Nine unconjugated bile acids, including the primary bile acids (cholic acid, chenodeoxycholic acid and α-muricholic acid) and the secondary bile acids (lithocholic acid, deoxycholic acid, ursodeoxycholic acid, hyodeoxycholic acid, β-muricholic acid and ω-muricholic acid) were quantified. Extraction conditions were evaluated, including: sample pH, type of organic solvent and amount of caecal material to be extracted. To compensate for sample matrix effects during extraction the method of standard addition was applied. The satisfactory linearity (r(2)>0.9840), high recovery (84.2-108.7%) and good intra-assay (6.3-10.6%) and inter-assay (6.9-11.1%) precision illustrated the good performance of the present method. The method is rapid, simple and capable of detecting and determining bile acids with limit of detection (LOD) ranged from 0.002 to 0.067μg/mL and limits of quantification (LOQ) varied from 0.006 to 0.224μg/mL. The results indicated that the concentration of some secondary bile acids, which usually are associated with health problems, were lower in rats fed with fermentable dietary fibre compared with a fibre free control diet, while the concentration of primary bile acids, usually connected with positive health effects, were higher in rats fed with diets containing dietary fibre. Of the dietary fibres, guar gum and to some extent the mixture of pectin+guar gum had the most positive effects. Thus, it was concluded that the composition of bile acids can be affected by the type of diet

Comprehensive on-line two-dimensional liquid chromatography × supercritical fluid chromatography with trapping column-assisted modulation for depolymerised lignin analysis

Lignin depolymerisation produces a large variety of low molecular weight phenolic compounds that can be upgraded to value-added chemicals. Detailed analysis of these complex depolymerisation mixtures is, however, hampered by the lack of resolving power of traditional analysis techniques. In this study, a novel online comprehensive two-dimensional reversed-phase liquid chromatography (RPLC) × supercritical fluid chromatography (SFC) method with trapping column interface was developed for the separation of phenolic compounds in depolymerised lignin samples. The trapping capacities of different trapping columns were evaluated. The influence of large volume water-containing injection on SFC performance was studied. The relation between peak capacity and first dimension flow rate and gradient was investigated. The optimized method was applied for the analysis of a depolymerised lignin sample. The RPLC × SFC system exhibited high degree of orthogonality. Compared with traditional loop based interface, trapping column interface can significantly shorten the analysis time and offer higher detectability, with the disadvantage of more severe undersampling in the first dimension

Signal enhancement in supercritical fluid chromatography-diode-array detection with multiple injection

To circumvent the detrimental effects of large-volume injection with fixed-loop injector in modern supercritical fluid chromatography, the feasibility of performing multiple injection was investigated. By accumulating analytes from a certain number of continual small-volume injections, compounds can be concentrated on the column head, and this leads to signal enhancement compared with a single injection. The signal to noise enhancement of different compounds appeared to be associated with their retention on different stationary phases and with type of sample diluent. The diethylamine column gave the best signal to noise enhancement when acetonitrile was used as sample diluent and the 2-picolylamine column showed the best overall performance with water as the sample diluent. The advantage of multiple injection over one-time large-volume injection was proven with sulfanilamide, with both acetonitrile and water as sample diluents. The multiple injection approach exhibited comparable within- and between-day precision of retention time and peak area with those of single injections. The potential of the multiple injection approach was demonstrated in the analysis of sulfanilamide-spiked honey extract and diclofenac-spiked ground water sample. The limitations of this approach were also discussed

A rapid method for analysis of fermentatively produced D-xylonate using ultra-high performance liquid chromatography and evaporative light scattering detection

An ultra-high performance liquid chromatogra-phy (UHPLC) based method for the analysis of D-xylonate was developed using an amide column in combination with an evaporative light scattering (ELS) detector. Separation of D-xylonate from other components of the fermentation medium was achieved. The dynamic range of the method was 0.2–7.0 g/L