Zeki Altun

Study of the factors affecting the performance of microextraction by packed sorbent (MEPS) using liquid scintillation counter and liquid chromatography-tandem mass spectrometry

Microextraction by packed sorbent (MEPS) is a new technique for sample preparation that can be connected on-line with LC or GC. In MEPS, approximately 1-2mg of the solid packing material is inserted into a syringe (100-250 microL) as a plug. Sample preparation takes place on the packed bed. The bed can be packed or coated to provide selective and suitable sampling conditions. The new method is very promising for extraction of drugs and metabolites from biological samples. In this paper, some factors affecting the performance of MEPS such as recovery, carry-over, leakage, washing volume and elution volume were studied using C18 and hydroxylated polystyrene-divinylbenzene copolymer (ENV+) as sorbents. Radioactively labelled bupivacaine in plasma samples was used as test analyte. For the extraction of this drug, using methanol/water 95:5 (v/v) (0.25% ammonium hydroxide) was used as elution solvent. The analyte response increased with increasing the elution volume and it was linear upp up to 100 microL utilizing liquid scintillation counter. Further, for concentrating the sample, we found that MEPS may be used such that the sample can be drawn through the needle, up and down, several times. The analyte leakage increases as the volume washing increases, though higher washing volumes may also result in cleaner extracts. To eliminate analyte carry-over, the sorbents were washed first with 3 x 250 microL elution solution and then with 3 x 250 microL washing solution. In addition, the reproducibility measurements show relatively good relative standard deviation (RSD) % values concerning analyte recovery and analyte leakage. The present study provides an understanding of basic aspects when optimizing methods for MEPS. In this study, MEPS was used off-line with liquid scintillation counter and on-line with LC-MS/MS.

Microextraction in Packed Syringe Online with Liquid Chromatography‐Tandem Mass Spectrometry: Molecularly Imprinted Polymer as Packing Material for MEPS in Selective Extraction of Ropivacaine from Plasma

Abstract The excellent performance of a new sample preparation method, microextraction in packed syringe (MEPS), was recently illustrated by online LC‐MS and GS‐MS assays of local anaesthetics in plasma samples. In the method, approximately 1 mg of solid packing material was inserted into a syringe (100–250 µL) as a plug. Sample preparation took place on the packed bed. The new method was easy to use, fully automated, of low cost, and rapid in comparison with previously used methods. This paper presents the use of molecularly imprinted polymers (MIPs) as packing material for higher extraction selectivity. Development and validation of a method for MIP‐MEPS online with LC‐MS‐MS using ropivacaine in plasma as model compound were investigated. A bupivacaine imprinted polymer was used. The method was validated and the standard curves were evaluated by means of quadratic regression and weighted by inverse of the concentration: 1/x for the calibration range 2–2000 nM. The applied polymer could be used more than 100 times before the syringe was discarded. The extraction recovery was 60%. The results showed high correlation coefficients (R2>0.999) for all runs. The accuracy, given as a percentage deviation from the nominal concentration values, ranged from −6% to 3%. The precision, given as the relative standard deviation, at three different concentrations (QC samples) was consistently about 3% to 10%. The limit of quantification was 2 nM.

Monolithic methacrylate packed 96-tips for high throughput bioanalysis.

In the pharmaceutical industry the growing number of samples to be analyzed requires high throughput and fully automated analytical techniques. Commonly used sample-preparation methods are solid-phase extraction (SPE), liquid-liquid extraction (LLE) and protein precipitation. In this paper we will discus a new sample-preparation technique based on SPE for high throughput drug extraction developed and used by our group. This new sample-preparation method is based on monolithic methacrylate polymer as packing sorbent for 96-tip robotic device. Using this device a 96-well plate could be handled in 2-4min. The key aspect of the monolithic phase is that monolithic material can offer both good binding capacity and low back-pressure properties compared to e.g. silica phases. The present paper presents the successful application of monolithic 96-tips and LC-MS/MS by the sample preparation of busulphan, rescovitine, metoprolol, pindolol and local anaesthetics from human plasma samples and cyklophosphamid from mice blood samples.

Increasing Sample Preparation Throughput Using Monolithic Methacrylate Polymer as Packing Material for 96-Tip Robotic Device

Abstract In this work, a laboratory robot has been used to accomplish a system for high sample cleanup throughput. The robot operating in a 96‐well format was furnished with 96 polypropylene tips packed with a chemically bonded monolithic methacrylate plug as sample adsorbent. Using this system, 96 samples could be handled in 2 minutes. Polypropylene tips were furnished with a chemically bonded monolithic methacrylate plug as sample adsorbent. Roscovitine and lidocaine in plasma samples were used as model substances. The validation of the methodology showed that the accuracy values of quality control samples (QC) were between +15%, and precision had a maximum deviation of 11%. The standard curve was obtained within the concentration range 14‐5600 nM in both plasma and water samples. The regression correlation coefficients (R2) for plasma and water samples were ≥0.999 for all runs. This paper was presented at HPLC 2005.

Evaluation of monolithic packed 96-tips and liquid chromatography–tandem mass spectrometry for extraction and quantification of pindolol and metoprolol in human plasma samples

96-well pipette tips with a chemically bonded monolithic methacrylate sorbent plug were used for solid-phase extraction (SPE) of pindolol and metoprolol in human plasma samples. The sorbent plug was formed by in situ polymerization. Monolithic packed 96-tips are a tool for miniaturized, solid-phase extraction. Using such packed 96-tips, a 96-well plate could be handled in about 2 min. The key aspect of the monolithic phase is that monolithic material can provide both relatively good binding capacity and relatively low backpressure properties. The validation of the methodology showed that the accuracy values of quality-control samples were between 101% and 103% for metoprolol, while between 94% and 114% for pindolol. The precision ranged from 4% to 15%. The standard calibration curves were obtained within the concentration range 5-5000 nM in plasma samples. The coefficients of determination (R2) for plasma samples were >or=0.99. Our prepared polymer based monolithic packed 96-tips were compared with commercial silica based 96-tips and protein precipitation.

Evaluation of monolithic packed 96-tips for solid-phase extraction of local anesthetics from human plasma for quantitation by liquid chromatography tandem mass spectrometry

Abstract Polypropylene 96‐well tips packed with a chemically bonded monolithic methacrylate plug as sample adsorbent were used for solid‐phase extraction (SPE) of ropivacaine and bupivacaine in human plasma samples. Monolithic packed 96‐tips are a miniaturized, solid-phase extraction. Packed 96‐tips require some microliter volumes of solvent (50–100 µL) for elution. Using packed 96‐tips, a 96‐well plate could be handled in about 2 minutes. The key aspect of the monolithic phase is that monolithic material processes both high binding capacity and low back‐pressure properties. The validation of the methodology showed that the accuracy values of quality control samples were between 101 and 118%, while the precision ranged from 4% to 17%. The standard calibration curves were obtained within the concentration range 2–2000 nM in plasma samples. The coefficients of determination (R2) for plasma samples were between 0.984 and 0.999.