Continuous fraction collection of gas chromatographic separations with parallel mass spectrometric detection applied to cell-based bioactivity analysis.

We describe the development and evaluation of a GC-MS fractionation platform that combines high-resolution fraction collection of full chromatograms with parallel MS detection. A y-split at the column divides the effluent towards the MS detector and towards an inverted y-piece where vaporized trap solvent is infused. The latter flow is directed outside the GC oven allowing subsequent condensation and stepwise collection of liquid fractions with trapped analytes on a 384-well plate. For study and optimization of the effluent split ratio, restriction capillaries of different lengths and diameters were evaluated. For a wide range of settings, local pressures were monitored during fractionation to assess the influence of MS vacuum and trap solvent infusion on the GC system stability. The platform performance was evaluated by GC-MS analysis and continuous fractionation of an n-alkane mixture followed by GC analysis of each fraction. Comparison of the on-line recorded and fraction-reconstructed chromatogram showed the GC separation is maintained during fractionation. Multiple fractionation cycles of the n-alkane sample on the same 384-well plate yielded a reconstructed chromatogram which was highly similar to that of a single analysis, demonstrating the high repeatability. The applicability of the GC-MS-fractionation platform for bioactivity screening was investigated by applying the AR-Ecoscreen reporter gene bioassay on fractions obtained after analysis of standard solutions and dust samples containing the anti-androgenic compounds vinclozolin and p,p'-DDE.

Gas chromatography fractionation platform featuring parallel flame-ionization detection and continuous high-resolution analyte collection in 384-well plates.

Gas chromatography (GC) is a superior separation technique for many compounds. However, fractionation of a GC eluate for analyte isolation and/or post-column off-line analysis is not straightforward, and existing platforms are limited in the number of fractions that can be collected. Moreover, aerosol formation may cause serious analyte losses. Previously, our group has developed a platform that resolved these limitations of GC fractionation by post-column infusion of a trap solvent prior to continuous small-volume fraction collection in a 96-wells plate (Pieke et al., 2013 [17]). Still, this GC fractionation set-up lacked a chemical detector for the on-line recording of chromatograms, and the introduction of trap solvent resulted in extensive peak broadening for late-eluting compounds. This paper reports advancements to the fractionation platform allowing flame ionization detection (FID) parallel to high-resolution collection of a full GC chromatograms in up to 384 nanofractions of 7s each. To this end, a post-column split was incorporated which directs part of the eluate towards FID. Furthermore, a solvent heating device was developed for stable delivery of preheated/vaporized trap solvent, which significantly reduced band broadening by post-column infusion. In order to achieve optimal analyte trapping, several solvents were tested at different flow rates. The repeatability of the optimized GC fraction collection process was assessed demonstrating the possibility of up-concentration of isolated analytes by repetitive analyses of the same sample. The feasibility of the improved GC fractionation platform for bioactivity screening of toxic compounds was studied by the analysis of a mixture of test pesticides, which after fractionation were subjected to a post-column acetylcholinesterase (AChE) assay. Fractions showing AChE inhibition could be unambiguously correlated with peaks from the parallel-recorded FID chromatogram.