Large-Ring Cyclodextrins as Chiral Selectors for Enantiomeric Pharmaceuticals.

Large-ring cyclodextrins (CD) are cyclic glucans composed of 9 or more α-1,4-linked glucose units. They are minor side products of bacterial glucanotransferases (CGTases, EC 2.4.1.19) and have previously been available only in very small amounts for studies of their properties in supramolecular complex formation reactions. We engineered a CGTase to synthesize mainly large-ring CD facilitating their preparation in larger amounts. By reversed phase chromatography, we obtained single CD samples composed of 10 to 12 glucose units (CD10, CD11, and CD12) with a purity of >90 %. Their identity was confirmed by high resolution mass spectrometry and fragmentation analysis. We demonstrated the non-toxicity of CD10-CD12 for human cell lines by a cell proliferation assay and impedimetric monitoring. We then showed that CD10 and CD11 are efficient chiral selectors for the capillary electrophoretic separation of the enantiomeric pharmaceuticals fluvastatin, mefloquine, carvedilol, and primaquine.

HPLC-MS with Glass Chips Featuring Monolithically Integrated Electrospray Emitters of Different Geometries.

We present and evaluate an approach for coupling liquid chromatography in glass chips with mass spectrometry via fully integrated electrospray emitters. We developed an instrumental platform which allows a robust and reproducible operation of high performance chip chromatography coupled to mass spectrometry. A comparison of differently shaped emitters, from flat over edged to pulled geometries, revealed that all types performed equally well for typical nano-HPLC flow rates. At very low flow rates below 50 nL·min(-1) very sharp, pulled nanospray emitters turned out to be mandatory for the generation of a stable electrospray.

Chip-based high-performance liquid chromatography for high-speed enantioseparations.

In this work, the first high-performance chiral liquid chromatography in packed microfluidic chips is presented. The chromatographic separation was performed on a column integrated into the microfluidic glass chip and packed with the particulate chiral stationary phase. Cellulose tris(3,5-dimethylphenylcarbamate) coated on 5-µm fully porous silica was used as chiral stationary phase material. Several racemic analytes including pharmaceutical products were baseline separated into their corresponding enantiomers under reversed-phase, polar organic and normal-phase conditions, demonstrating the versatility of the glass chip in the field of chiral separations. Van Deemter plots revealed a reduced plate height of 2.2 and a trend to enhanced mass transfer processes for solutes under low retention conditions. The utilization of very short column lengths of down to 12 mm led to ultrafast separations of enantiomers within 5 s.

High-performance liquid chromatography on glass chips using precisely defined porous polymer monoliths as particle retaining elements.

A stable and permanent integration of miniature packed bed separation columns into microfluidic systems is a major issue in nano liquid chromatography. Various approaches like differently shaped retaining elements or the use of key stone effect have been investigated. We show a flexible integration of miniature packed bed separation columns into microfluidic chips utilising common HPLC material achieved by laser-assisted generation of narrow, photopolymerised frits. The generated retaining elements serve as an in- and outlet frits for the columns. An optimised pre-polymeric solution, consisting of butyl acrylates and a porogen, allows a precise fabrication of frit-type structures with lengths of less than 100 m and the capability to withstand common slurry packing pressures of more than 250 bar. The separation of seven polycyclic aromatic hydrocarbons by pressure-driven, reversed-phase chromatography proves the high quality of the created chromatographic column inside a glass chip. Plate heights down to 2.9 were achieved and extremely fast separations with sub-second peak widths were performed in isocratic and gradient elution modes on very short columns (≤ 25 mm).