Rendering A Chiral Screening Step In Supercritical Fluid Chromatography Mass-Spectrometry Compatible.

Nowadays, sensitive chiral methods are required for the determination of chiral impurities and for assays in biological samples. Supercritical fluid chromatography (SFC), one of the main techniques to separate chiral molecules, can be coupled to MS to provide such sensitive methods. Moreover, chiral separation strategies are very useful to reduce the development time and cost of such methods. This study investigates the transfer of an existing non-MS compatible screening step (as part of a separation strategy) into an MS-compatible one. The initial step had a cumulative success rate of 100 % for 57 tested compounds using methanol or 2-propanol as mobile phase modifier on one of four chiral stationary phases. The additives applied in the original mobile phases, i.e. isopropylamine and trifluoroacetic acid, negatively affect the ionization in SFC-MS and thus need to be replaced. Formic acid, acetic acid, water, ammonia, ammonium acetate and ammonium formate were investigated as MS-compatible additives in different combinations and concentrations. Only methanol-based mobile phases were considered in this study because high system pressures were obtained with isopropanol. The other experimental parameters remained the same as in the initial screening step. The effects of the alternative additives on the obtained resolutions as well as on the global success rate were investigated. The best alternative MS-compatible mobile phase contained 0.5 % CH3COOH and 40 mM NH3 as additives. This mobile phase provided the highest number of separations and rather high resolutions. An MS-compatible screening step was defined with this alternative mobile phase. Compared to the original additives, a similar success rate was obtained.

CE-MS metabolic profiling of volume-restricted plasma samples from an acute mouse model for epileptic seizures to discover potentially involved metabolomic features.

Currently, a high variety of analytical techniques to perform metabolomics is available. One of these techniques is capillary electrophoresis coupled to mass spectrometry (CE-MS), which has emerged as a rather strong analytical technique for profiling polar and charged compounds. This work aims to discover with CE-MS potential metabolic consequences of evoked seizures in plasma by using a 6Hz acute corneal seizure mouse model. CE-MS is an appealing technique because of its capability to handle very small sample volumes, such as the 10 µL plasma samples obtained using capillary microsampling in this study. After liquid-liquid extraction, the samples were analyzed with CE-MS using low-pH separation conditions, followed by data analysis and biomarker identification. Both electrically induced seizures showed decreased values of methionine, lysine, glycine, phenylalanine, citrulline, 3-methyladenine and histidine in mice plasma. However, a second provoked seizure, 13 days later, showed a less pronounced decrease of the mean concentrations of these plasma metabolites, demonstrated by higher fold change ratios. Other obtained markers that can be related to seizure activities based on literature data, are isoleucine, serine, proline, tryptophan, alanine, arginine, valine and asparagine. Most amino acids showed relatively stable plasma concentrations between the basal levels (Time point 1) and after the 13-day wash-out period (Time point 3), which suggests its effectiveness. Overall, this work clearly demonstrated the possibility of profiling metabolite consequences related to seizure activities of an intrinsically low amount of body fluid using CE-MS. It would be useful to investigate and validate, in the future, the known and unknown metabolites in different animal models as well as in humans.

Defining a system suitability limit to decide on column deterioration and to facilitate column transfers in chiral supercritical fluid chromatography.

The separation of enantiomers is an important requirement during the entire drug life cycle in the pharmaceutical industry. High-performance liquid chromatography and supercritical fluid chromatography (SFC) are the main chromatographic techniques used to separate enantiomers. Since chiral stationary phases are often extensively used once a method has been developed, columns will age and must be replaced after a certain period. However, no practical guidelines exist to determine when a column is deteriorated or to decide whether a transfer to another column (with the same chiral selector) is successful. In this study, a system suitability limit for resolution was defined, based on an intermediate (time-different) precision study in SFC on four immobilized polysaccharide-based columns that only differed in manufacturer or particle size. This system suitability limit could be used to decide on column deterioration or as a requirement to evaluate whether a separation transfer was successful. Some method adaptations may be necessary to obtain successful transfers. An approach was proposed, which helped the analyst to make successful transfers. Graphical abstract.

Analytical techniques for metabolomic studies: a review.

Metabolomics is the comprehensive study of small-molecule metabolites. Obtaining a wide coverage of the metabolome is challenging because of the broad range of physicochemical properties of the small molecules. To study the compounds of interest spectroscopic (NMR), spectrometric (MS) and separation techniques (LC, GC, supercritical fluid chromatography, CE) are used. The choice for a given technique is influenced by the sample matrix, the concentration and properties of the metabolites, and the amount of sample. This review discusses the most commonly used analytical techniques for metabolomic studies, including their advantages, drawbacks and some applications.

Enantioseparations of pharmaceuticals with capillary electrochromatography: A review.

The chiral separation of pharmaceuticals is one of the major research topics in the pharmaceutical industry. Chromatographic techniques are most frequently used in this context. Separations in capillary electrochromatography (CEC) are an alternative and achieved by chromatographic retention and electrophoretic mobility principles. As a result, CEC is characterized by a high selectivity and efficiency. The limited number of stationary phases specifically developed for CEC, the low number of commercially available CEC columns, the frits to maintain the stationary phase, which forms fragile spots in the columns, and the limited column robustness and reproducibility, make CEC not very attractive for industrial application. However, CEC is still applied and studied in the academic field. This review discusses the enantioseparation of drugs in CEC published during the last four years, with a critical view on the reproducibility and the practical utility of these applications.