Biocompatible C18-polyacrylonitrile (PAN) coating was used as the extraction phase for an automated 96-blade solid phase microextraction (SPME) system with thin-film geometry. Three different methods of coating preparation (dipping, brush painting, and spraying) were evaluated; the spraying method was optimum in terms of its stability and reusability. The high-throughput sample preparation was achieved by using a robotic autosampler that enabled simultaneous preparation of 96 samples in 96-well-plate format. The increased volume of the extraction phase of the C18-PAN thin film coating resulted in significant enhancement in the extraction recovery when compared with that of the C18-PAN rod fibers. Various factors, such as reusability, reproducibility, pH stability, and reliability of the coating were evaluated. The results showed that the C18-PAN 96-blade SPME coating presented good extraction recovery, long-term reusability, good reproducibility, and biocompatibility. The limits of detection and quantitation were in the ranges of 0.1-0.3 and 0.5-1 ng/mL for all four analytes.

The main objective of metabolomics is the analysis of all lowmolecular-weight compounds present in a particular living system. Metabolomics data is complementary to proteomics, genomics, and transcriptomics data and provides a better understanding of dynamic processes occurring in living systems.[1] The processes of sampling and sample preparation can significantly affect the composition of the measured metabolome, so the analytical results may not adequately reflect the true metabolome composition at the time of sampling. [2][3][4] This is due primarily to poor efficiency (or even complete omission) of metabolism quenching step and multistep handling procedures, which contribute to inadvertent metabolite loss and/or degradation.Herein we introduce in vivo solid-phase microextraction (SPME) as a new sample preparation method for global metabolomics studies of living systems using liquid chromatography-mass spectrometry (LC-MS). SPME is a nonexhaustive sample preparation procedure in which the amount of analyte extracted is governed by the distribution coefficient of the analyte between the SPME coating and sample matrix if the equilibrium is reached or the rate of mass transfer if a short sampling time is used.[5] In vivo SPME allows accurate extraction of the metabolome directly in the tissue or blood of freely moving animals without the need to withdraw a representative biological sample for analysis, under conditions of negligible depletion where the amount of analyte extracted by SPME is independent of the sample volume. [5][6][7] The blood-draw-free nature of the sampling method facilitates multiple sampling of the same living system and can capture unstable or short-lived metabolites.Large biomolecules are not extracted by the specially selected biocompatible SPME coating, so the need for a metabolism quenching step is eliminated. The amount of metabolites extracted is proportional to the biologically active unbound concentration. For metabolomics studies, in vivo SPME provides the simplest and most rapid sample preparation tool available to date to study living systems in a format directly compatible with LC-MS detection. Although SPME was successfully applied to metabolomics studies using GC-MS primarily in headspace mode, [8][9][10][11][12] its capability to provide instantaneous metabolism quenching directly during the sampling process to capture true metabolome of blood or tissue has not been previously evaluated.First, we developed a successful in vivo SPME workflow for direct sampling of metabolome, and applied it to mice as a model system (Figure 1). In this approach, a coated SPME fiber is housed inside hypodermic needle, [13] which is used to pierce the sampling interface containing circulating blood. The fiber is exposed to blood for a pre-set short sampling time of 2 min. During the sampling, analytes are extracted directly into the SPME coating. The key aspect of developing SPME device for metabolomics was selection of the chemical nature of the coating to ensure simultaneous extraction of both...

Trigonal tricationic ionic liquids (ILs) are a new class of ILs that appear to be unique when used as gas chromatographic stationary phases. They consist of four core structures; (1) A = mesitylene core, (2) B = benzene core, (3) C = triethylamine core, and (4) D = tri(2-hexanamido)ethylamine core; to which three identical imidazolium or phosphonium cationic moieties were attached. These were coated on fused silica capillaries, and their gas chromatographic properties were evaluated. They were characterized using a linear solvation parameter model and a number of test mixtures. On the basis of the literature, it is known that both monocationic and dicationic ILs possess almost identical polarities, solvation characteristics, and chromatographic selectivities. However, some of the trigonal tricationic ILs were quite different. The different solvation parameters and higher apparent polarities appear to generate from the more rigid trigonal geometry of these ILs, as well as their ability to retain the positive charges in relatively close proximity to one another in some cases. Their unique selectivities, retention behaviors, and separation efficiencies were demonstrated using the Grob mixture, a flavor and fragrance test mixture, alcohols/alkanes test, and FAME isomer separations. Two ILs C1 (methylimidazolium substitution) and C4 (2-hydroxyethylimidazolium substitution) had higher apparent polarities than any know IL (mono, di, and tricationic ILs) or commercial stationary phases. The tri(2-hexanamido)ethylamine core IL series proved to be very interesting in that it not only showed the highest separation efficiency for all test mixtures, but it also is the first IL stationary phase (containing NTf(2)(-) anions) that eliminates peak tailing for alcohols and other H-bonding analytes. The thermal stabilities were investigated using three methods: thermogravimetric analysis (TGA) method, temperature programmed gas chromatographic method (TPGC), and isothermal gas chromatographic method. The D core series had a high working temperature range, exceptional selectivities, and higher separation efficiencies than comparable polarity commercial columns. It appears that this specific type of multifunctional ILs may have the most promising future as a new generation of gas chromatographic stationary phases.

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Four new ionic liquids (IL) were prepared and bonded onto 5-microm silica particles for use as adsorbent in solid-phase microextraction (SPME). Two ILs contained styrene units that allowed for polymerization and higher carbon content of the bonded silica particles. Two polymeric ILs differing by their anion were used to prepare two SPME fibers that were used in both headspace and immersion extractions and compared to commercial fibers. In both sets of experiments, ethyl acetate was used as an internal standard to take into account adsorbent volume differences between the fibers. The polymeric IL fibers are very efficient in headspace SPME for short-chain alcohols. Immersion SPME also can be used with the IL fibers for short-chain alcohols as well as for polar and basic amines that can be extracted at pH 11 without damage to the IL-bonded silica particles. The sensitivities of the two IL fibers differing by the anion were similar. Their efficacy compares favorably to that of commercial fibers for polar analytes. The mechanical strength and durability of the polymeric IL fibers were excellent.