On-line coupling of RPLC and SFC for the analysis of chiral compounds

https://www.ilmexhibitions.com/htc/abstract/On-line+coupling+of+RPLC+and+chiral+SFC+for+the+analysis+of+pharmaceutical+compounds/378/International audienc

On-line achiral RPLCx chiral SFC for the analysis of chiral compounds

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Comprehensive two dimensional liquid chromatography as analytical strategy for pharmaceutical analysis

International audienceComprehensive on-line two-dimensional liquid chromatography (LCxLC) is expected to generate impressive peak capacities, which makes it a method of choice for the analysis of complex samples such as pharmaceuticals. A comparative study of different sets of chromatographic conditions including stationary phase, pH additive and organic modifier was carried out with two real pharmaceutical samples in order to find out the best analytical conditions for implementation of one or several generic on-line LCxLC separations. Our choice was based on the evaluation of both degree of orthogonality and practical sample peak capacity under linear gradient conditions. The potential of 190 combinations of chromatographic systems was compared. A set of 3 RPLCxRPLC configurations was found to be very attractive for both samples and in good agreement with the findings of a previous study carried out with 17 model compounds, thereby supporting the idea of using generic LCxLC conditions in the pharmaceutical area. The three selected 2D-systems were implemented for the on-line RPLCxRPLC-UV/MS analysis of two pharmaceutical samples. It was shown, for each sample, that these 2D-systems were able to generate an effective peak capacity close to 1000 in less than 50min. For each sample, baseline separation was obtained for every known compound and furthermore a large number of unknown impurities could also be separated and identified. Finally, in the proposed conditions, the total number of compounds detected was significantly improved from one RPLC separation to one RPLCxRPLC separation. Only a small additional gain was observed by performing a second RPLCxRPLC separation or even a third one

Characterization of positional isomers of drug intermediates by off-line RPLC x SFC hyphenated to high resolution MS

The authors greatly acknowledge Agilent Technologies for the loan of the q-TOF instrument.International audienceMany steps are needed in the synthesis of a new active pharmaceutical ingredient (API). In a practical case proposed by a French pharmaceutical company, an intermediate synthesis step, needed to protect 8 hydroxyl groups before oxidation, could produce a mixture of neutral compounds containing up to 652 structures being positional isomers of 18 molecular formulas. Some mixtures allowed obtaining the desired API, others did not. An efficient analytical method was needed to characterize these neutral positional isomers and identify the mixtures to reject. Two samples were provided by the pharmaceutical company: Sample A was conform, Sample B was not. 8 RPLC columns were used with different gradients to screen Sample A. Next, the best RPLC separation was used as the second dimension fast analysis in a comprehensive 2D-RPLC systems. Two columns were used as first dimension: a fluorinated one and a zirconium based one. An order of magnitude was gained in peak capacity, but a better sample characterization was still needed. An off-line RPLC x SFC x Q-TOF/MS analysis was performed collecting 96 RPLC fractions and analyzing them by SFC with Q-TOF/MS detection. A home-made software associated the 96 SFC MS chromatograms to produce either base peak (BPC) or extract ion (EIC) contour plots that allowed for a satisfying characterization of the samples. Subtracting the EIC of expected m/z compounds from the Sample B BPC contour plot produced a unique new contour plot clearly pointing out unexpected compounds explaining the failure of the synthesis and possibly allowing improving the synthesis process

Vegetative storage proteins in white clover (Trifolium repens L.) : quantitative and qualitative features

International audienceThe kinetic pattern of protein mobilization in roots, stolons and nodules of white clover (Trifolium repens cv. Grasslands Huia) was studied over a regrowth period following complete defoliation. Defoliation led to a significant decrease in soluble protein in stolons and roots during the first days of regrowth as compared with uncut plants, Protein degradation was also observed in nodules of both uncut and defoliated plants. Two of the proteins characterized (15 and 17.3 kDa), which accumulate mainly in perennial tissues. have previously been referred to as Vegetative Storage Proteins (VSPs). Using plants inoculated with either efficient (potentially functional) or inefficient Rhizobium strains, the 15 kDa VSP appeared to be located exclusively in the nodules. and it cross-reacted positively with antibodies raised against soybean leghaemoglobin. Nevertheless, the kinetics of its hydrolysis-accumulation following defoliation clearly supported the view that it may play a role in nitrogen storage. The 17.3 kDa protein was shown to accumulate in response to exposure to low temperature. and exhibited a seasonal pattern of relative accumulation under field conditions. Results are discussed in terms of the putative role that this VSP may play in overwintering of clover

Collaborative study of a Supercritical Fluid Chromatography method for the determination of pharmaceutical impurities

Projet PHAR

First inter-laboratory study of a Supercritical Fluid Chromatography method for the determination of pharmaceutical impurities

International audienceSupercritical Fluid Chromatography (SFC) has known a strong regain of interest for the last 10 years, especially in the field of pharmaceutical analysis. Besides the development and validation of the SFC method in one individual laboratory, it is also important to demonstrate its applicability and transferability to various laboratories around the world. Therefore, an inter-laboratory study was conducted and published for the first time in SFC, to assess method reproducibility, and evaluate whether this chromatographic technique could become a reference method for quality control (QC) laboratories. This study involved 19 participating laboratories from 4 continents and 9 different countries. It included 5 academic groups, 3 demonstration laboratories at analytical instrument companies, 10 pharmaceutical companies and 1 food company. In the initial analysis of the study results, consistencies within- and between-laboratories were deeply examined. In the subsequent analysis, the method reproducibility was estimated taking into account variances in replicates, between-days and between-laboratories. The results obtained were compared with the literature values for liquid chromatography (LC) in the context of impurities determination. Repeatability and reproducibility variances were found to be similar or better than those described for LC methods, and highlighted the adequacy of the SFC method for QC analyses. The results demonstrated the excellent and robust quantitative performance of SFC. Consequently, this complementary technique is recognized on equal merit to other chromatographic techniques