Mirlinda Biba

Systematic evaluation of new chiral stationary phases for supercritical fluid chromatography using a standard racemate library.

A systematic approach to the evaluation of new chiral stationary phases (CSPs) for supercritical fluid chromatography (SFC) using a standard library of racemic analytes is described. A standard library of racemic analytes representing a variety of functional group classes was assembled from a mixture of proprietary and commercial compounds. The library is dispensed and stored in a convenient 96-well microplate format to facilitate ease of use, and to minimize the amount of analyte required for analysis. Automated SFC screening was performed on both established CSPs in common use, as well as a group of six recently commercialized CSPs. Screening results were archived in a structure-searchable database that allows convenient comparison of performance data to determine which CSPs shows the best performance.

Comparison of Multiparallel Microfluidic HPLC Instruments for High Throughput Analyses in Support of Pharmaceutical Process Research

Abstract The Eksigent Express 800 8 channel and the Nanostream Veloce 24 channel microfluidic HPLC systems were compared for their utility in providing high throughput chromatographic analysis to support a growing demand in pharmaceutical process research. Criteria for success are discussed, a comparison of the two instruments is made, and results from a head to head evaluation using a variety of different analyte mixtures are presented.

Improved chiral SFC screening for analytical method development.

In this study we describe the evaluation of a recently developed supercritical fluid chromatography (SFC) instrument for automated chiral SFC method development. The greatly improved gradient dwell volume and liquid flow control of the new instrument in combination with the use of shorter columns containing smaller stationary phase particles affords chiral SFC method development that is faster and more universal than previous systems.

Effect of particle size on the speed and resolution of chiral separations using supercritical fluid chromatography.

Fast chiral supercritical fluid chromatography (SFC) separations have become important due to the increasing use of high-throughput experimentation (HTE) in organic synthesis. These HTE experiments can generate hundreds of samples for chiral analysis that need to be assayed in a short time. In general, chiral SFC can provide much faster analysis times compared to liquid chromatography (LC). Additionally, columns packed with smaller particles can provide faster and more efficient separations. In this study, the effect of the particle size on the speed and resolution of chiral separations by SFC was evaluated. The performance of Chiralcel OD columns packed with either 5 or 3 μm particles were compared using van Deemter or other kinetic plots. The benefits of using smaller particle columns for chiral SFC analysis are illustrated.

Microplate evaluation of process adsorbents

A microplate-based assay to rapidly identify adsorbents suitable for removing process impurities is described. A solution containing both product and impurity is added to a number of wells, each containing a small amount of a candidate adsorbent. After equilibration, analysis of the supernatant solution allows one to determine the extent of adsorption of both product and impurity. Fast analysis techniques such as flow injection analysis LC-MS or the use of colorimetric indicators allows rapid identification of the most selective adsorbents for a given separation problem.

Factors influencing the separation of oligonucleotides using reversed-phase/ion-exchange mixed-mode high performance liquid chromatography columns

New mixed-mode columns consisting of reversed-phase and ion-exchange separation modes were evaluated for the analysis of short RNA oligonucleotides (∼20mers). Conventional analysis for these samples typically involves using two complementary methods: strong anion-exchange liquid chromatography (SAX-LC) for separation based on charge, and ion-pair reversed-phase liquid chromatography (IP-RPLC) for separation based on hydrophobicity. Recently introduced mixed-mode high performance liquid chromatography (HPLC) columns combine both reversed-phase and ion-exchange modes, potentially offering a simpler analysis by combining the benefits of both separation modes into a single method. Analysis of a variety of RNA oligonucleotide samples using three different mixed-mode stationary phases showed some distinct benefits for oligonucleotide separation and analysis. When using these mixed-mode columns with typical IP-RPLC mobile phase conditions, such as ammonium acetate or triethylammonium acetate as the primary ion-pair reagent, the separation was mainly based on the IP-RPLC mode. However, when changing the mobile phase conditions to those more typical for SAX-LC, such as salt gradients with NaCl or NaBr, very different separation patterns were observed due to mixed-mode interactions. In addition, the Scherzo SW-C18 and SM-C18 columns with sodium chloride or sodium bromide salt gradients also showed significant improvements in peak shape.

A chiral 1,4-oxazin-2-one: asymmetric synthesis versus resolution, structure, conformation and VCD absolute configuration

Abstract 1,4-Oxazin-2-one 3 is obtained from 2-pinanone in 4 steps and 78% overall yield. Enantiopure (e.e. >99%) (R)-(+)-3 and (S)-(−)-3 were obtained through chiral supercritical fluid chromatography (using a semi preparative Chiralpak AS column) with almost quantitative recovery of material. The structure and the boat-conformation of the lactone ring have been determined by NMR and the absolute configuration determined by VCD.

Serendipitous discovery of a pH-dependant atropisomer bond rotation: Toward a write-protectable chiral molecular switch?

Owing to slow rotation of a sterically constrained dimethylamide substituent, two slowly interconverting enantiomers of a preclinical candidate for pharmaceutical development, 1, (6-(3-Chloro-4-fluoro-benzyl)-4-hydroxy-2-methyl-3,5-dioxo-2,3,5,6,7,8-hexahydro-[2,6]naphthyridine-1-carboxylic acid dimethylamide) are observed by chiral chromatography. Isolation of pure enantiomer by preparative chiral chromatography followed by enantiopurity analysis over time allowed for a study of the kinetics of enantiomer interconversion under a variety of conditions. Relatively slow racemization was observed in alcohol solvents, with a half life on the order of 5-10 h. A dramatic influence of aqueous buffer pH on racemization was noted, with higher pH leading to rapid racemization within a few minutes, and lower pH leading to essentially no racemization for periods up to a week. A hypothesis explaining this unusual effect of pH on carboxamide bond rotation is offered, and some suggestions for potential utility of such a system are considered.

Factors influencing the interconversion of a new class of dibenzodiazepine sulfonamide atropisomers

A novel family of atropisomers based on a conformationally constrained seven membered ring system is investigated using a combination of preparative chiral chromatography, circular dichroism, and other analytical techniques. The influence of structure on the rate of atropisomer interconversion was explored with a series of analogs showing a range of interconversion rates ranging from very fast (undetectable on the HPLC timescale) to very slow (half life of many days).

Evaluation of core-shell particle columns for ion-pair reversed-phase liquid chromatography analysis of oligonucleotides.

An investigation into the use of core-shell particle columns for separation of short (∼21 base pairs) RNA oligonucleotides by ion-pair reversed-phase liquid chromatography (IP-RPLC) showed improved resolution for a number of test analytes relative to conventional (fully-porous) reversed-phase columns. The best resolutions were obtained using columns packed with smaller sub-2μm core-shell particles.