Compared to conventional one-dimensional gas chromatography (1D-GC), comprehensive two-dimensional gas chromatography (GCÃ-GC) offers increased peak capacity, improved resolution and enhanced mass sensitivity. In addition, it generates structured two-dimensional (2-D) chromatograms, which aids in the identification of compound classes. Sample preparation procedures can often be minimized, or even eliminated in some cases, due to the superior separating power offered by the technique. All of these advantages make GCÃ-GC a very powerful tool in environmental analysis involving the determination of trace levels of toxic compounds in complex matrices. This review paper summarizes and examines some of the GCÃ-GC applications in environmental analysis and monitoring.
Keywords Comprehensive two-dimensional gas chromatography (GCÃ-GC) . Environmental analysis . Water and sediment . PCBs/PCDDs/PCDFs . Pesticides . Air analysis
Many years of industrialization and urbanization have resulted in the worldwide distribution of numerous chemicals throughout the atmosphere, hydrosphere and lithosphere. Many of these compounds can be hazardous to the world's ecosystems and to humans. Environmental analytical chemists have the task of analyzing these compounds in the environment. Whenever the analytes have reasonably high vapor pressures, gas chromatography is the method of choice.
The main challenge in environmental analysis is that the analytes are usually present in trace amounts in very complex matrices. As a result, tremendous research effort goes into the analysis of major environmental pollutants, including polychlorinated dibenzodioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), polychlorinated biphenyls (PCBs), polycyclic aromatic hydrocarbons (PAHs), organochlorines, pesticides and endocrine disruptors [1].
The approach utilized in environmental analysis is generally the same as in any analytical procedure. It consists of sampling, sample preparation, separation and detection. All of these steps could benefit from improvements, yet it is usually the separation step that imposes the biggest limitations. In the case of gas chromatography (GC), most environmental samples contain so many closely eluting peaks of analytes and matrix components that altogether the peak capacity in one chromatographic dimension (1D) is greatly exceeded, and numerous coelutions and/or entire unresolved regions are observed during the separation. This leads to poor identification and quantification of the analytes of interest [1].
Poor chromatographic resolution places high demands on both sample preparation and detection instrumentation. Sample preparation can be expensive and labor-intensive, and can generate large amounts of environmentally harmful solvent waste. The development of microextraction approaches, such as liquid-liquid microextraction (LLME) and solid-phase microextraction (SPME), and solvent-free sample introduction systems (e.g., direct thermal desorption), has the potential to vastly simplify the sample preparation process without sacrificing the sensitivity and/or selectivity [2-5]. With respect to detection, insufficient GC resolution usually means that the use of mass spectrometry (MS), including high-resolution mass spectrometry (HRMS) in some cases, is mandatory. Figure 1 illustrates the common problem encountered in GC-MS [36]. In a 1D-GC analysis of a food extract spiked with pesticides (Fig. 1b), trace amounts of the analytes of interest (in this case chlorfenvinphos) commonly coelute with more abundant components of the sample matrix. As a result, the mass spectra obtained for such compounds (Fig. 1e) frequently contain fragments originating from the interfering compounds, causing poor matching with the library mass spectra (Fig. 1d). MS deconvolution algorithms can significantly improve the quality of spectral information for coeluting peaks, but they are not always successful when the number of coelutions is high. As shown in Fig. 1a, comprehensive 2D-GC (GCÃ-GC) increases the separation space and improves the chromatographic resolution, leading to separation of the analyte of interest (chlorfenvinphos) from the coeluting compounds and/or matrix components. As a result, the quality of the mass spectra of the analytes is improved (Fig. 1c), allowing for more confident analyte identification (Fig. 1d). It is still possible that some coelutions will be present; these can usually be efficiently resolved with MS deconvolution, which produces better results when the number of coeluting components is reduced. The increased separation power of GCÃ-GC thus leads to successful analyte identification and/or quantification.
Fig. 1a-e GCÃ-GC-TOF MS versus 1D-GC-TOF MS for the analysis of a carrot extract. a GCÃ-GC-TOF MS contour plot; b 1D-GC-TOF MS chromatogram of the same region: upper trace, TIC scaled to 1%; lower trace, m/z 323 ion trace; c Mass spectrum obtained after GCÃ-GC separation showing the characteristic m/z values of chlorfenvinphos (m/z 81, 109, 267, 295, 323). d Library spectrum of chlorfenvinphos and e spectrum obtained at the retention time of chlorfenvinphos after 1D-GC separation [36].
Historically, the peak capacity problem in conventional gas chromatography was dealt with through the implementation of multidimensional gas chromatography (MDGC). In this method, also referred to as "heart-cutting," a complex and unresolved portion of a one-dimensional (1D) chromatogram is subjected to additional separation on a second column coated with a stationary phase of different selectivity [1]. Although this approach does increase the resolution of the selected portion(s) of the 1D chromatogram, automation of this method is challenging, and only some sample components can be fully resolved. Nevertheless, numerous applications dealing with the analysis of PCBs, pesticides and toxaphene, among others, have been reported with varying degrees of success [6-12]. Overall, however, it is clear that many separations would benefit if the entire sample was subjected to separation in two dimensions. This became possible with the introduction of comprehensive two-dimensional gas chromatography (GCÃ-GC).
Comprehensive Two-Dimensional Gas Chromatography (GCÃ-GC)
Comprehensive 2D-GC is the ultimate solution to the peak capacity problem encountered in 1D-GC. A block diagram of a typical GCÃ-GC set-up is illustrated in Fig. 2. Most instrumental components utilized in GCÃ-GC are in fact the same as in 1D-GC. These include the injector, the oven, the columns and the detectors. In a typical GCÃ-GC system, a long column coated with a thick film of a nonpolar stationary phase is installed as the primary column. Its outlet is connected through a special interface, or modulator, to the inlet of a second dimension column, coated with a stationary phase of different selectivity. The modulator not only physically connects the primary and the secondary columns; its main role is to repeatedly trap the components of the effluent from the first dimension and periodically inject them in the form of narrow pulses into the secondary column for further chromatographic separation. Since the second dimension operates under fast GC conditions, detector choices in GCÃ-GC are limited to those capable of fast data acquisition rates. Examples of detectors that were found suitable for GCÃ-GC include a flame ionization detector (FID), an electron capture detector (ECD), an atomic emission detector (AED), a sulfur chemiluminescence detector (SCD), a nitrogen chemiluminescence detector (NCD), and a time-of-flight mass spectrometer (TOF MS).
Fig. 2 A block diagram of a GCÃ-GC system. a injector; b primary column; c modulator or interface; d secondary column; and e detector
The modulator is the heart of the instrument because it ensures that the separation is both comprehensive (the entire sample is subjected to both separation dimensions) and multidimensional (separation accomplished in one dimension is not lost in the other dimension) [13]. Since the first practical implementation of GCÃ-GC in 1991, the field has witnessed numerous modulator designs [14]. Initially, thermal modulators utilizing heating to modulate were implemented; however, modulation with cryogenic liquids (liquid CO2 or N2) is presently predominant. Within the cryogenic modulator family, each design has its own distinct advantages and limitations, making it suitable for specific types of analyses. For example, an interface for the analysis of surface water contaminants was developed [19], while the construction and application of an in-house modulator for the quantitative analysis of PAHs and PCBs was described [15]. Shortly thereafter, different types of modulators in the analysis of organohalogenated analytes was evaluated [16].
The implementation of GCÃ-GC offers the following advantages over 1D separation methods: enhanced separation power; improved mass sensitivity (observed only with thermal modulators due to the focusing effect); and structured, or highly ordered, chromatograms. In environmental analysis, GCÃ-GC has the potential to improve separation of the toxic compounds from the coeluting analytes and matrix components, to increase the detection limits of such chemicals, and to provide structured two-dimensional chromatograms ideal for monitoring applications. Consequently, this can lead to minimized sample preparation procedures, and hence decreased analysis time. Other applications are also possible. For example, most recently, GCÃ-GC has been used for the estimation of environmental partitioning properties of diesel fuel hydrocarbons, which are important in oil spills affecting many ecosystems [17].
GCÃ-GC in Environmental Analysis and Monitoring
Water and Sediment Analysis
Drinking water is the most essential substance to life on this planet. In order to assess the safety of potable water for humans, there is a need for rapid, precise and accurate methods for its monitoring and analysis. River and lake sediments also play an important role in the health of aquatic ecosystems; therefore, they must be analyzed as well. Typical methods for the analysis of water pollutants include time-consuming sample preparation, followed by GC-MS analysis. It was realized early on that GCÃ-GC has a great potential to improve the analysis of water and sediment pollutants.
In one of its earliest applications in this area, GCÃ-GC was illustrated can fully separate the BTEX fraction (benzene, toluene, ethyl benzene and xylenes) and methyl tert-butyl ether (MTBE) from common matrix interferences in a single run when combined with SPME [18]. The improved separation power of GCÃ-GC, both MTBE and benzene were baseline-resolved in the 2-D chromatographic space. This study illustrated the great potential of GCÃ-GC used in combination with a microextraction technique (headspace SPME) for rapid identification and monitoring of aqueous pollutants.
Some of the Earth's freshwater bodies are subject to daily petroleum and oil contamination from numerous sources. As a result, there is a high demand for the quick and reliable analysis of water and sediments from the affected sites. In the 1970s, it was noticed that chromatograms of petroleum samples exhibited a characteristic, unresolved, raised baseline "hump" [20]. This complex part of the chromatogram, consisting of many different classes of compounds, is presently referred to as an "unresolved complex mixture" (UCM) [21]. GCÃ-GC-FID was used to resolve the UCM in two different freshwater sediments [21]. Adhering to conventional sample preparation procedures, the authors took advantage of the superior resolving power and structured chromatograms generated by GCÃ-GC to study the different fractions of the sediment's UCM. The chromatograms obtained for both samples provided the researchers with clues pointing to the main source of contamination. More importantly, the research illustrated the potential of GCÃ-GC in environmental forensics, an essential tool for environmental chemistry, environmental law and environmental audits.
In the 1990s, it was realized that surfactants such as nonylphenol (NP) isomers, degradation products of nonylphenol polyethoxylate, are possible estrogen disruptors [22]. Adding to the concern, NPs are presently found in water and sediments from urban areas [23]. GCÃ-GC-TOF MS was used for the separation of NP isomers from a technical mixture [24]. Forty-one components were identified. Figure 3 illustrates the application of GCÃ-GC-TOF MS in the analysis of individual ion traces of NP isomers from the same study. Two NP fragmentation products are illustrated, m/z 135 (Fig. 3a) and m/z 149 (Fig. 3b). Both chromatograms exhibit group-type separations, emphasized by sloped lines connecting peak maxima of compounds within the same homologous family. It is evident from this figure that because of the structural similarity of various NP isomers, complete separation is very difficult. Nevertheless, the additional resolution power provided by GCÃ-GC provided "cleaner" mass spectra, which made analyte identification much easier.
Fig. 3a, b Extracted ion GCÃ-GC-TOF MS chromatograms of a technical nonylphenyl (NP) mixture; a m/z 135; b m/z 149. The sloping lines connect peak maxima for a homologous series (i.e. m/z 135 and m/z 149). Both chromatograms illustrate 6 different lines (solid and dashed), indicating the presence of 6 different families of NP isomers
GCÃ-GC was recently applied for the analysis of environmental pollutants present in marine sediments [25]. Morales-Munoz et al. developed a method for qualitative, fast and high-resolution analysis of complex samples based on dynamic ultrasound-assisted extraction (UAE) coupled to GCÃ-GC-TOF MS. UAE offers a fast and efficient alternative to sample pretreatment procedures applied to solid samples [30]. A combination of an efficient and selective sample preparation method, UAE, with a powerful separation method, GCÃ-GC, led to the resolution of 1500 compounds and identification of several polycyclic aromatic hydrocarbons (PAHs), NPs and dialkylated benzenes. Once again, the ability of GCÃ-GC to not only separate analytes from each other but also from the sample matrix proved invaluable.
Suspected carcinogens and mutagens, PAHs are byproducts of many industrial activities and are ubiquitously distributed in the environment. Their trace determination in sediment samples is difficult because it requires laborious and selective sample preparation. In an effort to improve trace analysis of PAHs in complex matrices, Cavagnino et al. utilized a large-volume splitless injection (LVSI) technique in conjunction with GCÃ-GC-FID [26]. The complexity of the samples analyzed was representative of many sediment samples obtained from rivers and lakes. Separation and detection of seven PAHs diluted in composite diesel fuel at low ppb levels demonstrated the potential of LVSI- GCÃ-GC-FID as a powerful and rapid tool for the analysis of trace amounts of PAHs in complex matrices.
Around the same time, Ong et al. developed a method for rapid monitoring of PAHs in soil samples, utilizing pressurized liquid extraction (PLE)-GCÃ-GC-FID [27]. The currently published work is merely an indicator of the potential applicability of GCÃ-GC in the analysis of PAH in sediment samples.
Overall, with improved resolution, enhanced mass sensitivity and ordered chromatograms, GCÃ-GC can be coupled with effective and rapid sample preparation methods to yield results unachievable by conventional analytical procedures.
Analysis of PCBs, PCDDs and PCDFs
Polychlorinated dibenzodioxins (PCDDs), polychlorinated dibenzofurans (PCDFs) and some polychlorinated biphenyl (PCB) congeners are subject to bioaccumulation and biomagnification in the environment and are thus dangerous to wildlife and humans. Many of them are suspected carcinogens and mutagens [28]. Confident assessment of PCBs, dioxins and furans in the environment requires a method capable of isolating and quantifying them in complex samples such as foods, soil and water.
GCÃ-GC offers an advantageous approach to the analysis of PCBs, PCDDs and PCDFs in complex matrices. In one of its early experiments, a liquid crystal primary column (separation based on planarity) and a nonpolar secondary column (separation based on vapor pressure) were used to separate mono - and non-ortho PCB congeners from a technical mixture [30]. GCÃ-GC linked to microelectron-capture detection (MECD) was applied for the determination of toxic PCBs, PCDDs and PCDFs in cod liver samples [29]. The results of the analysis illustrated full separation and identification of all 12 priority PCB congeners as well as the most toxic dioxins and furans from liver samples spiked with 90 PCBs and 17 toxic PCDDs and PCDFs. Additionally, when compared to standard sample preparation procedures, the liver sample pretreatment was nonselective and minimized. It consisted of cell lysis, centrifugation and fractionation followed by direct injection into the GCÃ-GC system. Figure 4 illustrates the 2-D chromatogram obtained from the analysis of the cod liver sample. Most recently, a multilaboratory study was performed, analyzing PCDD/Fs and WHO PCBs in food samples by comparing GCÃ-GC-MECD with GC-HRMS, and illustrating once again the great potential of GCÃ-GC in rapid monitoring applications [31].
In an effort to compare GCÃ-GC to standard methods of analysis, the performance of GCÃ-GC coupled to 13C-label isotope dilution (ID) TOF MS (GCÃ-GC-ID-TOF MS) with conventional GC-HRMS was evaluated [32]. Quantification of 17 PCDD/Fs and four PCBs spiked onto soil and sediment samples was comparable for both methods. However, GCÃ-GC implementation required only minimal sample preparation and resulted in signal enhancement (factor of 5-10), superior resolution, lower instrumentation costs, and improved spectral deconvolution of the TOF MS data [32]. As a consequence of the increased peak capacity and resolution, identification of unknown compounds was possible.
Fig. 4 GCÃ-GC-ECD chromatogram of a cod liver sample spiked with 90 PCBs [29].
Pesticide Analysis
Analysis of pesticides poses challenges to analytical chemists with respect to both sample preparation procedures and chromatography. Similar to other toxic compounds, pesticides are usually distributed throughout the environment in trace amounts. Additionally, they are part of extremely complex matrices such as food, soil and water samples. The need for rapid high-resolution methods of analysis is as pressing today as it ever was.
Early applications of GCÃ-GC to pesticide analysis in human tissues illustrated the potential of the method for routine implementation in the future. Supercritical fluid extraction (SFE) in conjunction with GCÃ-GC-FID was utilized to analyze pesticides in human serum [33]. The baseline separation of 15 pesticides extracted from spiked human serum was achieved in less than four minutes. Later, GCÃ-GC-FID was utilized for assessing pesticide exposure of children by using small volumes of urine and serum [34]. This particular example illustrated complete separation of 16 pesticides in less than four minutes. More recently, the identification and quantification of 59 tissue contaminants was demonstrated which including PCBs and organochlorine pesticides [35]. The authors indicated that for such an analysis, three separate injections are required in standard routine analyses (GC-ID-TOFMS), while GCÃ-GC-ID-TOF MS accomplished comparable results in a single run. Such approaches should prove invaluable in clinical settings, where a relatively cheap, reliable and reproducible method is needed for screening and positive rapid identification of target analytes.
Determination of pesticides in food extracts is equally important. Utilizing GCÃ-GC-TOF MS to separate and identify all 58 pesticides spiked onto vegetables was illustrated [36]. This was accomplished with minimal and nonselective sample preparation: a celery or carrot sample was chopped, mixed with sodium acetate and ethyl acetate, blended, centrifuged and dried. The extract was injected into the GCÃ-GC [36]. It should be noted that although not all 58 pesticides were baseline-resolved, the high resolving power of the GCÃ-GC method allowed proper identification of partially coeluting peaks through the use of TOF MS spectral deconvolution algorithms. The results of deconvolution are usually not as good in conventional GC-TOF MS experiments, as the method lacks the resolving power to chromatographically separate trace pesticides from the surrounding and abundant matrix components.
Recently, five different GCÃ-GC column combinations for the group separation of 12 halogenated compound classes was evaluated, which consisting of PCBs, PCDDs, PCDFs, polychlorinated diphenyl esters (PCDEs), polychlorinated naphthalenes (PCNs), polychlorinated dibenzothiophenes (PCDTs), polychlorinated terphenyls (PCTs), polychlorinated alkanes (PCAs), toxaphene, polybrominated biphenyls (PBBs), polybrominated diphenyl ethers (PBDEs) and organochlorine pesticides (OCPs) [37]. Although the focus of this paper was primarily group separation of different compound classes, separations within families were also illustrated. While the separation and identification of all 28 OCPs was demonstrated only for a pure pesticide mixture, most of them were fully separated even when injected alongside the other eleven compound classes [37]. Hence, it seems that with a correctly configured column set, GCÃ-GC can be used as a primary screening step for environmental samples contaminated with pesticides alongside many other classes of pollutants, with minimal sample preparation.
Regardless of the pesticide type or the complexity of the sample, GCÃ-GC provides a more powerful tool for rapid analysis than the presently utilized 1D methods. Increased resolution and decreased analysis time make it perfectly suitable for monitoring purposes.
Air Analysis
Volatile organic compounds (VOCs) play an important role in the generation of urban photochemical smog [38]. The World Health Organization (WHO) recognizes that exposure to air particulate matter can have detrimental effects on human health [39]. Still, uncertainty exists with regard to the health effects from VOCs in urban particulate matter (PM) [40]. Thus, a rapid, reliable and informative method is required to ensure successful monitoring, identification and discovery of atmospheric pollutants.
Many PAHs and oxygenated PAHs (oxy-PAHs) are suspected carcinogens and mutagens, and so they are important target analytes in urban aerosol analysis. GCÃ-GC-FID and GCÃ-GC-quadrupole MS (QMS) applied to urban air samples from Finland allowed the detection of approximately 1500 peaks and identification of target PAHs [41]. The GCÃ-GC-FID combination confirmed good reproducibility, while the MS-coupled approach was used for compound identification and quantification. Thirteen non-target PAHs were identified, and ten target PAHs were quantified. The PAH concentration range found (0.5-5.5 ng/m3) was comparable to results obtained by standard methods in other parts of Europe [41].
The suitability of direct thermal desorption combined with GCÃ-GC (DTDGCÃ-GC-TOF MS) was investigated for the analysis of organic compounds in ambient aerosol particles [42]. The use of DTD as a sample introduction method eliminated solvent use. When compared to similar analysis with GC-TOF MS, GCÃ-GC-TOF MS exhibited a ten-fold increase in the number of peaks detected and produced highly structured chromatograms ideal for rapid screening purposes. More importantly, the comprehensive 2D-GC approach reduced the limitations of TOF MS deconvolution observed in 1D; this led to improved library matches and more confident analyte identification.
Cigarette smoke is an extremely complex mixture estimated to consist of approximately 4,700 identified compounds and up to 100,000 unidentified components [43]. GCÃ-GC-TOFMS was utilized to resolve approximately 30,000 peaks from cigarette smoke [43]. Following this, simpler samples of cigarette smoke condensate were analyzed to determine the chemical composition of the neutral fraction [44], the basic fraction [45], and the acidic fraction [46]. Conventional GC-MS can separate 200 unknown peaks and identify 115 hydrocarbons from the nonpolar neutral fraction of cigarette condensate; GCÃ-GC analysis of the same sample, however, achieved the separation of 4,000 compounds and the identification of 1,800 hydrocarbons [44]. In another study, GCÃ-GC-TOF MS analysis of the basic fraction of cigarette condensates identified 377 nitrogen-containing compounds, among which 155 were pyridine derivatives, 104 quinoline/isoquinoline derivatives and 56 pyrazine derivatives [45].
Conclusions
GCÃ-GC has quickly achieved the status of being one of the most powerful tools for the analysis of volatile organic compounds. It has established itself as a technology that is perfectly suitable for monitoring analytes in complex samples. In the area of environmental analysis, this is evidenced by the numerous examples of analysis of common environmental pollutants-including PCBs, PCDDs, PCDFs, PAHs and pesticides-in complex environmental matrices. Additionally, GCÃ-GC has the potential to simplify the sample preparation procedures (or even eliminate them entirely), while simultaneously generating high-resolution chromatograms in a shorter overall analysis time.
For a new analytical method to be widely adopted, it must not only be reliable and reproducible, but it should also exhibit significant advantages over accepted methods. The examples reported in this review illustrate the advantages of the GCÃ-GC method over traditional 1D-GC separations. During the first years in GCÃ-GC history, instrumentation development was the main focus; however, since the commercialization of GCÃ-GC systems, the number of reported applications in environmental analysis and other scientific fields have increased dramatically. Hence, we can expect a gradual transition to automated GCÃ-GC coupled to on-line sample preparation devices for applications in routine environmental monitoring.
Completed Date: 2011-04-23
