Preparation and use of packed capillary columns in chromatographic and related techniques

Self Cite

A column switching LC method is presented for the analysis of fluoxetine (FLU) and norfluoxetine (NFLU) by direct injection of human plasma using a lab-made restricted access media (RAM) column. A RAM-BSA-octadecyl silica (C-18) column (40 mm x 4.6 mm, 10 microm) is evaluated in both backflush and foreflush elution modes and coupled with a C-18 lab-made (50 mm x 4.6 mm, 3 microm) analytical column in order to perform online sample preparation. Direct injection of 100 microL of plasma samples is possible with the developed approach. In addition, reduction of sample handling is obtained when compared with traditional liquid-liquid extraction (LLE) and SPE. The total analysis time is around 20 min. A LOQ of 15 ng/mL is achieved in a concentration range of 15-500 ng/mL, allowing the therapeutic drug monitoring of clinical samples. The precision values achieved are lower than 15% for all the evaluated points with adequate recovery and accuracy. Furthermore, no matrix interferences are found in the analysis and the proposed method shows to be an adequate alternative for analysis of FLU in plasma.

Section: Column Preparationmentioning

confidence: 99%

Fluoxetine and norfluoxetine analysis by direct injection of human plasma in a column switching liquid chromatographic system

Santos-Neto

Fernandes

Rodrígues

et al. 2008

Self Cite

“…Over the last thirty years, many different methods have been introduced for the manufacture of capillary columns containing microparticulate stationary phases [15]. Slurry packing with a high pressure pump [16,17] is probably the most widely used strategy, although centrifugal force [18,19], gravity [20], vacuum [21] and electrokinetic [22] packing methods have also been utilized.…”

Section: Introductionmentioning

confidence: 99%

Capillary action liquid chromatography

Zhang

Bergström

Goodall³

et al. 2009

Capillary action LC (caLC) is introduced as a technique using capillary action as the driving force to perform LC in capillary columns packed with HPLC type microparticulate materials. A dry packing method with centrifugal force was developed to prepare capillary columns in parallel (10 columns per 3 min) to support their disposable use in caLC. Using a digital microscope for real-time imaging and recording separations of components in a dye mixture, caLC was found to have flow characteristics similar to TLC. Based on the investigation of microparticulate HPLC silica gels of different size (1.5-10 microm) and a typical TLC grade irregular medium, Merck 60G silica, the van Deemter curves suggested molecular diffusion as the major contribution to band broadening in caLC. With Waters Xbridge 2.6 microm silica, plate heights down to 8.8 microm were obtained, comparable to those achievable in HPLC. Assisted by an image-processing method, the visual caLC separation was converted to a classical chromatogram for further data analysis and such a facility confirmed the observation of highly efficient bands.

“…E-mail: christian.huber@mx.uni-saarland.de. mass transfer of solutes into and out of the stagnant mobile phase present in porous separation media and the need for devices that retain the particles in the capillary tube [13], representing an additional source of band broadening and/or irreversible analyte adsorption [14,15]. An alternative approach to the use of very small and/ or totally nonporous particles to alleviate the problem of mass transfer and intraparticular void volume is the concept of monolithic chromatographic beds.…”

Section: Introductionmentioning

confidence: 99%

Comparing monolithic and microparticular capillary columns for the separation and analysis of peptide mixtures by liquid chromatography‐mass spectrometry

Toll

Wintringer

Schweiger-Hufnagel

et al. 2005

A mixture of ten proteins was trypsinized and injected onto poly-(styrene-divinylben-zene) monolithic columns (60 x 0.20 or 0.10 mm ID) and a column packed with C18 silica particles (75 x 0.075 mm ID), respectively. The columns were eluted at 200-2000 nL/min with gradients of ACN in 0.050% TFA. Eluting peptides were detected by ESI-MS/MS and subsequently identified by database searching. The 100 microm ID monolithic column showed the highest cumulative Mowse scores based on the highest ion scores for the peptides and the largest number of identified peptides. It is shown that the number of identified peptides strongly depends on the dynamic range within the peptide mixture. In consequence, all proteins were identified in a mixture of relatively balanced analyte amounts (12.5-80 fmol) whereas only peptides for six out of ten proteins were found in a sample of high-dynamic range (0.65-270 fmol). The 100 microm monolithic column showed the highest reproducibility for peptide identifications in three consecutive runs. Depending on sample amount, 57-72% of the identified peptides were detectable in each of the three runs of triplicate analyses. The results demonstrate the high suitability of 100 microm monolithic columns for high-resolution peptide separations in proteomic research.