4 May / June 2014
isoprenoids mono- di- and tri aromatics, tri- and pentacyclic terpanes, steranes and aromatic steranes), in these cases, it is often necessary the introduction of an additional procedure during sample preparation. This step consists of the oil’s fractionation into saturated hydrocarbons, aromatics, resins, and asphaltenes (i.e., SARA separation) [7,8]. The biomarkers of interest are mostly found in the saturated fraction and, therefore, only the fraction of interest is subjected to GC- MS analysis [1]. During GC-MS analysis, the detection of the analytes is performed under full scan (for untargeted analysis) or single ion monitoring (SIM) mode (for targeted analysis). In either detection mode, under carefully devised experiments, the required qualitative information is obtained and used to provide the necessary information for chemical characterisation of the sample. Nonetheless, the results obtained by these methods are potentially jeopardised by co-elution. For instance, an ion with m/z 217 corresponds to a key fragment pertaining to steranes by electron ionisation. However, other molecules may exhibit this ion into its fractionation and co-elution with interfering molecules that exhibit similar fragmentation patterns (i.e., also exhibit m/z 217) will lead to erroneous interpretation of the chromatographic data [7,8,10]. If prior knowledge of the biomarkers is available [11], additional selectivity is obtained by exploring gas chromatography coupled to a tandem mass spectrometer (GC-MSn
).
For instance, multiple reaction monitoring (MRM) experiments are used exclusively for targeted analysis and/or structural elucidation [1,12,13].
3. Multidimensional Gas Chromatography (MDGC)
Multidimensional separation techniques are powerful methods in which two or more independent separative techniques are linked together for separation [14]. Multidimensional gas chromatographic (MDGC) techniques comprise of two or more independent gas chromatographic separations (i.e, dimensions) coupled in a sequential fashion [15,16]. The paramount requirement to achieve higher peak capacity, (maximum number of peaks separable), is that all GC stationary phases examined must possess distinct/complementary solvation capabilities and, thus, different selectivities [17]. Also, MDGC experiments must be devised to maintain, at least in part, the separation achieved in each dimension so that the resolving power of the composite separation exceeds that of the individual stages [16,17]. While this section is largely confined to the applications of MDGC techniques to the chemical characterisation of crude oils, several excellent reviews describing the fundamentals of MDGC are available for the interested reader [16,18,19- 23].
The use of MDGC techniques for the
analysis of biomarkers in petrochemical samples has yielded promising results [7,8,11,12]. These techniques can be divided into non-comprehensive and comprehensive methods. The former are heart-cutting MDGC techniques that employ valve-based or flow-controlled microfluidic devices, being the latter collectively known as Deans switch interfaces [19,20]. In these experiments, selected heart-cuts (i.e., fractions of the eluent of a previous dimension) are sampled and transferred to another GC column for additional separation. Therefore, this system allow that only a fraction of the entire sample experiences two or more GC separations, independent of the dimensions of the second column and the sample rate. Although these methods exhibit optimum analytical performance for targeted analysis, the overall increase in peak capacity is modest compared to comprehensive setups [24], which are recommended for untargeted analysis or discovery-based approaches.
Comprehensive MDGC techniques are those in which the whole sample, or a representative fraction of the sample, experience two or more GC separations [24-26]. This is accomplished by means of an interface that continuously samples and periodically transfers the chromatographic band to another dimension for additional separation. These interfaces can be: (a) valve-based, where the focusing of the analytes is effectuated by very rapidly sampling only very narrow fractions from the first column, however, about 90% of the effluent is vented off, so that the far less then 10% of the original injected amount of the sample is transferred to the second column, for this reason, this modulator can does not provide a comprehensive separation, being the type of modulator less used among all [14], (b) flow-controlled, this interface also uses pneumatic valves, here the effluent from the first dimension is accumulated in a loop and then be transferred to the second dimension, however, a loss of considerable percentage of effluent from the first column
can take place, in addition this device is subject to providing band broadening, however, new developments are using ultra rapid flow in the second dimension in order to avoid the loss of analyte transferred. [14,23] or (c) thermal modulators, in which accumulated of the analytes in a thick film capillary (modulation capillary) by thermal means (i.e. liquid nitrogen, carbon dioxide or dry ice) and its remobilisation it performed by applicating heat, resulting in narrows peaks and a increase of the peak amplitude and consequently are the type modulator more widely used [14,25]. They operated at sufficiently high sampling frequencies to preserve the separation achieved in the previous dimension.
Is worth noting that, if the peak capacity in the first and second column are, respectively, n1
and n2 +n2 peaks, the total peak capacity in
a GC-GC system will increase arithmetically for n1
peaks (remembering that, only a
fraction of the sample of the first dimension is transferred to the second dimension), while that in a system GC×GC, where the sample is subjected to separation by two columns of different selectivities, the peak of capacity will increase geometrically for n1
×n2 , for this reason, the theoretical
increase in peak capacity is far greater than those exhibited by heart-cutting techniques [15]. Introduced in 1991 by late Prof. John Phillips [27], comprehensive two-dimensional gas chromatography (GC×GC) is considered by many separation scientists as the biggest milestone in chromatography since the development of GC capillary columns in the late 50’s. One of the most important hallmarks of GC×GC is the presence of structured chromatograms where a relationship between the analytes molecular structure and retention coordinates is readily observable. For instance, paraffins, tricyclic terpanes, steranes, and hopanes elute in well defined regions of the chromatogram (see Figure 1) when a saturated fraction of a Brazilian oil sample is analysed by GC×GC coupled to a mass spectrometer with a time
Figure 1. GC×GC-TOFMS chromatogram from the saturated fraction of oil (Camamu-Almada basin, Brazil). 1
D column: 30 m × 0.25 mm HP-5MS (polydimethyldiphenylsiloxane with 5% diphenylsiloxane monomer incorporation) (df
= 0.25 µm). 2 diphenylsiloxane monomer incorporation) (df = 0.10 µm) adapted from Aguiar et al. [10]. D column: 150 cm × 0.10 mm DB-17 (polydimethyldiphenylsiloxane with 50%
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