Search

The definition of a chromatographic analysis speed based simply on analysis time is an outdated concept to define conventional chromatography, fast chromatography, and emerging high-resolution techniques such as comprehensive two-dimensional and comprehensive three-dimensional gas chromatography. Here, the metric average theoretical peak time (ATPT) is proposed for separation speed, considering conventional and multidimensional separations. ATPT can be defined as the time (in ms per peak) needed to elute a theoretical peak in a chromatographic system. Using this metric, it is possible to define ranges, proposed for a normal speed (ATPT higher than 4000 ms/peak), high speed (ATPT range from 600 to 4000 ms/peak), very high speed (ATPT range from 200 to 600 ms/peak), hyper speed (ATPT range from 3.3 to 200 ms/peak) and ultra high speed chromatography (ATPT lower than 3.3 ms/peak), that combines time and efficiency metrics. This metric was applied in several contexts to demonstrate its robustness to evaluate chromatographic separations for different techniques and analytical conditions. Applications also demonstrate the advantages of the use of ATPT as a method development metric tool., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier B.V.)

The influence of the modifier concentration in supercritical mixtures of carbon dioxide and methanol on the speed-resolution properties of columns packed with 1.7μm core-shell particles was investigated from a theoretical viewpoint. Molar fractions of methanol up to 30% were considered. The column was assumed to be operated under strict adiabatic conditions in order to maximize its efficiency. Four inlet temperatures were tested, between 297 and 337K. Four different pressure drops along the column were considered, between 25 and 200bar. The physico-chemical properties (density, viscosity, and heat capacity) of the mixtures of carbon dioxide and methanol were derived from the NIST REFPROP program for temperatures between 287 and 337K and pressures between 150 and 390bar. The axial heterogeneity of the column was taken into account by segmenting it into 500 hundreds slices in each of which all physical and chromatographic properties were assumed to be uniform. The apparent kinetic Poppe plots were built from the apparent column efficiency calculated from the sum of the increments of the retention times and time variances from the column inlet to its outlet. The numerical results showed that the axial heterogeneity of the column due to axial variations of the temperature and the equilibrium constant decreases with increasing molar fraction of methanol in the eluent when the pressure drop is increased from 25 to 200bar. The methanol content decreases the speed-resolution of the column more particularly when the analysis is done at low pressure drops. The results demonstrate also that under adiabatic conditions, for pressure drops larger than 100bar, an increase of methanol concentration does not cause a dramatic loss of speed-resolution. For example, at 310K and with a pressure drop of 25bar, the longest column is expected to deliver 17,000 plates; increasing the molar fractions of methanol from 0 to 30% decreases this efficiency by about 15% and increases the analysis time by +140%. In contrast, under the same experimental conditions except for a pressure drop of 200bar, the maximum efficiency (130,000 plates) would remain unchanged and the analysis time increase by only 40%., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Background: A rapid liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed and validated for the quantification of reactive oxygen species (ROS) derived free oxysterols and cholesterol in human plasma and atherosclerotic plaque., Method: In vitro autoxidation of cholesterol during sample pretreatment was avoided by applying only one protein precipitation and re-concentration step using 80 μl plasma. For preparation of 10mg atherosclerotic plaques an additional liquid-liquid extraction was included. Free 7-keto-, 7-α/ß-hydroxy-, 5,6-α-epoxy-, 5,6-β-epoxycholesterol, cholestane-3ß,5α,6ß-triol and cholesterol were separated within 7 min on a monolithic column. An API 4000 tandem mass spectrometer was applied in positive ionization mode using atmospheric pressure chemical ionization., Results: The detection limit was 0.1 ng/ml and the linearity ranged from 0.5 to 0.75 to 2000 ng/ml for the oxysterols and from 50 to 1000 μg/ml for cholesterol. Recovery was between 80.9 and 107.9%. Between-run imprecision ranged from 7.9 to 11.7%. Analysis of plasma samples from additional 50 middle-aged volunteers revealed a large inter-individual variability (e.g. 7-ketocholesterol 2.63-30.47 ng/ml). Oxysterol concentrations normalized to cholesterol were about 43 times higher in carotid plaque compared to plasma (n=5)., Conclusion: This rapid LC-MS/MS method enables reliable quantification focused on especially ROS-derived oxysterols in human plasma and atherosclerotic plaque samples under high-throughput conditions., (© 2013. Published by Elsevier B.V. All rights reserved.)