Roger Trones

Temperature-programmed packed capillary liquid chromatography separation with large volume on-column focusing of retinyl esters

Summary A non-aqueous isocratic reversed-phase packed capillary high performance liquid chromatography method for the determination of retinyl esters, utilizing temperature programming and on-column focusing large volume injection, has been developed. The stationary phase material was C30, and the mobile phase consisted of acetonitrile-dichloromethane (70 : 30, v/v). A three-step temperature program, starting at 10 C for 10 min, then 1 /min to 30 C, and finally 2.5 /min to 70 C, was found most appropriate. Compared to an isothermal separation at 25 C, this temperature program provided improved peak resolution, enhanced peak shapes of the last eluting compounds, and a reduction of the overall elution time. A mass limit of detection of 27 pg was found with respect to retinyl palmitate, using UV detection with an “U” shaped flow cell at 327 nm. This corresponds to a concentration limit of detection of 2.7 pg/ L, when utilizing an injection volume of 10 L. The concentration of retinyl palmitate in arctic seal liver samples was estimated to be 62.6 g/g liver.

Development of a flat membrane based device for electromembrane extraction: a new approach for exhaustive extraction of basic drugs from human plasma.

In this work, a single-well electromembrane extraction (EME) device was developed based on a thin (100μm) and flat porous membrane of polypropylene supporting a liquid membrane. The new EME device was operated with a relatively large acceptor solution volume to promote a high recovery. Using this EME device, exhaustive extraction of the basic drugs quetiapine, citalopram, amitriptyline, methadone and sertraline was investigated from both acidified water samples and human plasma. The volume of acceptor solution, extraction time, and extraction voltage were found to be important factors for obtaining exhaustive extraction. 2-Nitrophenyl octyl ether was selected as the optimal organic solvent for the supported liquid membrane. From spiked acidified water samples (600μl), EME was carried out with 600μl of 20mM HCOOH as acceptor solution for 15min and with an extraction voltage of 250V. Under these conditions, extraction recoveries were in the range 89-112%. From human plasma samples (600μl), EME was carried out with 600μl of 20mM HCOOH as acceptor solution for 30min and with an extraction voltage of 300V. Under these conditions, extraction recoveries were in the range of 83-105%. When combined with LC-MS, the new EME device provided linearity in the range 10-1000ng/ml for all analytes (R(2)>0.990). The repeatability at low (10ng/ml), medium (100ng/ml), and high (1000ng/ml) concentration level for all five analytes were less than 10% (RSD). The limits of quantification (S/N=10) were found to be in the range 0.7-6.4ng/ml.

Microconcentric nebulizer for the coupling of micro liquid chromatography and capillary zone electrophoresis with inductively coupled plasma mass spectrometry

A concentric nebulizer designed to deliver less than 10 µl min -1 directly into an ICP-MS system is described. The influence of the liquid flow rate, the nebulizer gas flow rate and the distance from the nebulizer tip to the plasma was evaluated with regard to sensitivity and precision. The nebulizer was tested at liquid flow rates of 3–7 µl min -1 . Samples of tetramethyllead and tetraethyllead dissolved in dimethylformamide were used for the evaluation of the nebulizer. The detection limit for Pb ( m =208) with a direct injection system was found to be 0.3 pg (S/N=3) or 5 µg l -1 , based on an injection volume of 60 nl, a liquid flow rate of 5 µl min -1 and a dwell time of 10 ms. The precision was found to be 1.9% ( n =10) based on peak area measurements. The responses of tetramethyllead and tetraethyllead were not significantly different ( p =0.05). The time needed for one determination was less than 15 s, owing to the quick rinse out of the nebulizer. The nebulizer can be used as part of a direct injection instrumental set-up. An application of the nebulizer to the coupling of packed capillary liquid chromatography with ICP-MS detection is demonstrated.

Separation of polyethylene glycol oligomers using inverse temperature programming in packed capillary liquid chromatography

Inverse temperature programming in packed capillary liquid chromatography coupled to evaporative light-scattering detection has been used to resolve native polyethylene glycol (PEG) oligomers. The model compound, PEG 1000, was separated on a 300 mm x 0.32 mm I.D. capillary column packed with 3 microm Hypersil ODS particles with acetonitrile-water (30:70, v/v) as mobile phase. The retention of the PEG oligomers increased with increasing temperature, different from what is commonly observed in liquid chromatography. The retention times of the oligomers were approximately doubled for each 25 degrees C increment of the column temperature in the temperature range 30-80 degrees C. The oligomers were almost unretained and co-eluted at a column temperature of 30 degrees C. At 80 degrees C a baseline separation of more than 22 peaks was obtained, but the last eluting peaks were severely broadened and all oligomers did not elute. When a negatively sloped temperature ramp from 80 to 25 degrees C at -1.5 degrees C/min was applied, the peak shapes were improved, additional peaks were detected and the analysis time was reduced by 48%. In the temperature programming mode, the intra-day precision of the retention times ranged from 0.5 to 5.8% (n=5).

Aspects and applications of non-aqueous high temperature packed capillary liquid chromatography

The effect of using high isothermal temperatures and temperature programming above ambient in liquid chromatography has been investigated by the use of relatively long packed reversed phase capillary columns and non-aqueous mobile phases. Efficiency measurements have been performed in the temperature interval 50– 175 °C, indicating that the best efficiency was achieved at 100 °C, with a corresponding optimum linear velocity of approximately 0.07 cm s–1. The efficiency measurements revealed that the linear velocity had to be slightly increased in order to operate at the optimum reduced plate height at elevated temperatures. The contribution from extra-column volumes increased the plate height at temperatures above 100 °C where the solute had low retention. This effect was closely examined by introducing larger pre-column dead volumes, demonstrating the need to minimize dead volumes. Temperature programming has successfully been used for partial characterization and for purity testing of different polymer additives. The within and between day precision of retention times for the temperature-programmed separations gave a relative standard deviation of 0.3–2.9%.

Hindered amine stabilizers investigated by the use of packed capillary temperature-programmed liquid chromatography: I. Poly((6-((1,1,3,3-tetramethylbutyl)-amino)-1,3,5-triazine-2,4-diyl)(2,2,6,6-tetramethyl-4-piperidyl)imino)-1,6-hexanediyl((2,2,6,6-tetramethyl-4-piperidyl)imino))

Three different trademarks of a hindered amine stabilizer with the IUPAC name poly((6-((1,1,3,3-tetramethylbutyl)-amino)-1,3,5-triazine-2,4-d iyl)(2,2,6,6-tetramethyl-4-piperidyl)imino)-1,6-hexanediyl(( 2,2 ,6,6-tetramethyl-4-piperidyl)imino)), have been analyzed and compared to each other by the use of non-aqueous packed capillary temperature-programmed liquid chromatography and light scattering detection. The analysis by this method has shown that the products contained almost 40 different homologues and other components. This is in contrast to what has been assumed earlier based on results achieved with size exclusion chromatography. The method demonstrated significant differences between the products from different manufacturers.

Hindered amine stabilizers investigated by the use of packed capillary temperature programmed liquid chromatography: II: Poly-(N-β-hydroxyethyl-2,2,6,6-tetramethyl-4-hydroxypiperidyl succinate)☆

Three different trademark products of a hindered amine stabilizer with the IUPAC name poly-(N-beta-hydroxyethyl-2,2,6,6-tetramethyl-4-hydroxypiperidyl succinate), have been analyzed and compared to each other by the use of packed capillary temperature programmed liquid chromatography and light scattering detection. The analysis by this method has shown that the products contained approximately 50 different homologues and/or other components. The method also demonstrated its potential in purity control of products from different manufacturers.

Packed capillary high-temperature liquid chromatography coupled to inductively coupled plasma mass spectrometry

The applicability of inductively coupled plasma mass spectrometry (ICP-MS) as a detector for high-temperature liquid chromatography with packed capillary columns has been studied. A laboratory-made micro concentric nebulizer was utilized as a interface for the coupling of the two instruments. The repeatability of peak height and area at the investigated column temperatures (75–150°C) was good, with relative standard deviation less than 1.5% (n=3). The limit of detection (LOD) for tetraethyl- and tetramethyllead was found to be 5 pg as Pb (S/N=3), and for tetraethyltin 9 pg as Sn (S/N=3). The effect of temperature ramping on the baseline was investigated, and no limitations regarding range or rate of temperature increase upon the ICP-MS response were found for the mobile phases used.

Determination of polyethylene glycol in low-density polyethylene by large volume injection temperature gradient packed capillary liquid chromatography

Polyethylene glycol (PEG) 20000 in low-density polyethylene has been determined using column switching and inverse temperature programming in reversed-phase packed capillary liquid chromatography with evaporative light scattering detection. PEG 20000 was extracted into water from the polyethylene dissolved in toluene and PEG 35000 was added as an internal standard (I.S.). The samples in aliquots of 100 microl were reconcentrated on the enrichment column using a loading mobile phase of acetonitrile-water (3:97, v/v) at a flow-rate of 75 microl/min for 3 min, then back-flushed and separated on the analytical column with acetonitrile-THF-water (40:5:55, v/v) as mobile phase. The column temperature was reduced from 68 to 55 degrees C with a ramp of -1.5 degrees C/min, held constant for 3 min and then reduced further to 45 degrees C with a -1.5 degrees C/min ramp and kept constant for 1 min. The analysis runtime was 20 min. The recovery of PEG 20 000 was determined to 65.1% with 2.8% RSD and the mass limit of detection of PEG 20 000 was 1.25 microg. The within-assay and between day precision of the retention times of both PEG 20000 and PEG 35000 displayed RSD of less than 1.1% (n = 9), while the overall area ratio RSD of 100 microg/ml PEG 20000 over PEG 35000 was 1.3% (n = 9). The method was linear within the investigated concentration range 25-125 microg/ml (R2 = 0.9983). In addition, a mixture of PEG 4000, 8000, 10000, 20000 and 35000 was analysed on the system to demonstrate the possibility of analysing several PEGs in a sample with the use of temperature gradient elution.