Multidimensional gas chromatography with parallel cryogenic traps

than two parallel cryogenic traps and multiple analytical GC columns could be useful. INTRODUCTION. The combination of chromatography with spectroscop...
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Anal. Chern. 1993, 65,1012-1016

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Multidimensional Gas Chromatography with Parallel Cryogenic Traps N. Ragunathan, Kevin A. Krock, and Charles L. Wilkins' Department of Chemistry, University of California, Riverside, Riverside, California 92521

I n Its most common form, multidimensional gas chromatography (MDGC) has been restricted to use of a single Intermediate cryogenic trap between two serlally coupled columns. This paper descrlbes successful Implementationof a multi-port valve MDGC system with two parallel cryogenlc traps interposed between a first stage GC precolumn and a second stage GC analytlcal column. Using thls strategy, It Is possible to accommodatethe constraintsof infrared and mass spectrometry detectorsIn a directly llnkedstructure eiucidatlon system. Analysis of an unleaded gasoline sample serves to demonstrate the analytical advantages. Following a preliminary separation, uslng only the first column, a second portion of the same sample Is InJected for GC-GC analysis. Heart cuts from four different segments of the chromatogram identified in the preilminary Separation are analyzed uslng an analytlcal column of dlff erent selectlvlty. The prellmlnary slngle-stage Separation of the four chromatogram areas contained 7, 3, 12, and 12 peaks for cuts A, B, C, and D respectively, for a total of 34 peaks. The second stage separation of heart cuts A, B, C, and D yields 16, 17, 30, and 30 peaks, totaling 93 peaks. These results demonstrate significantly enhanced chromatographlc resolution uslng chromatographic conditlons permlttlng jolnt use of lnformatlonrlch infrared and mass spectral detectors. Thus, It Is suggested that MDGC systems employing more than two parallel cryogenic traps and multiple analytical GC columns could be useful.

INTRODUCT1 0N The combination of chromatography with spectroscopic detection provides a powerful method for analysis of complex multicomponent mixtures.',* A primary advantage of this approach derives from the fact that, in principle, chromatography can resolve or separate a multicomponent mixture into its individual constituents, and spectroscopic analysis of the separated substances can permit their unambiguous identification. However, in practice, complete chromatographic resolution of complex mixtures is rarely achieved by a single stage of analysis. This can be understood in terms of the concept of statistical column capacity. In their important study of this issue, Davis and Giddings3 used Poisson statistics to relate the expected practical capacity for randomly distributed chromatographic peaks to maximum column capacity, n,. They found that, for complex mixtures without any type of sample dilution (no reduction in m, the number of detectable components in a sample),the maximum ratio of p (the number of resolved peaks) to nc (column capacity) is 37 5%. Furthermore, the maximum ratio of single component peaks to column capacity (s/n,) is 18%. These results suggest that no more than 37% of the total column (1)Wilkins, C. L. Anal. Chem. 1987,59, 571A-581A. (2) Cooper, J. R.; Wilkins, C. L. Anal. Chem. 1989, 61, 1571-1577. (3) Davis, J . M.; Giddings, J. C. Anal. Chem. 1983, 55, 418-424.

capacity contains peaks and that no more than 18% of the capacity will be comprised of single-component peaks. The rest of the components will coelute to a greater or lesser degree. However, reducing the number of components or the dynamic range of a sample decreases the saturation value a , where a = m/n,, and, therefore, increases the probability of isolating a particular consituent. As a result of the constraints upon chromatography inherent in using a combined infrared and mass spectral detection scheme, the reliability of spectroscopic library search approaches or other analytical algorithms for qualitative analysis is necessarily compromised. Because use of these information-rich detectors precludes the use of the highest resolution chromatography methods, for single-stage chromatography, many of the measurements will be made on incompletely resolved mixtures of two or more components. Therefore, improved separation strategies are mandatory, if the true analytical potential of combined separation and multispectral analysis systems is to be realized. A partial solution to the problem of inseparability of a complex mixture by a single-stage separating apparatus is use of a multistage separation procedure. In such a multidimensional separation,mixtures being analyzed are subjected to two or more independent sequential separation steps.4For a capillary gas chromatography (GC) system, multidimensional separation is achieved by coupling two or more columns having different selectivity, with the provision that only a small chromatogram segment from the primary column is allowed to elute into the secondary column. This technique of analyzing small segments (or heart cuts) of a complex chromatogram by means of secondary analytical columns was first demonstrated by Simmons and Snyder in 1 9 X 5 The potential advantages of a multidimensional approach over a more conventional single-stage unidimensional system have been recognized and discussed thoroughly in many review articles.6-10Of course, for a great many applications, singlestage GC analysis is adequate. In general, applicationswhere a multidimensional system would be potentially most useful are for analysis of moderately or extremely complex mixtures such as, for example, hydrocarbon mixtures,ll essential oils,12 and environmental sarnples.l3 Two distinct modes of applying a multidimensional gas chromatographic (MDGC)method would be either for target (4) Giddings, J. C. Anal. Chem. 1984, 56, 1258A-1270A. (5) Simmons, M. C.; Snyder, L. R. Anal. Chem. 1958, 30, 32-35.

(6) Bertsch, W. ;I. High Resolut. Chromatogr. Chromatogr. Commun. 1978. 1. 85-90. Bertsch, W. J . High Resolut. Chromatogr.Chromatogr. Commun. I , 187-194. Bertsch, W. J . High Resolut. Chromatogr.Chromatogr. Commun. 1, 289-299. Himberg, H.; Sippola, E.: Riekkola, M. L. J . Microcol. Sep. 1989, 1 , 271-275. ( I 0 ) Bertsch, W. In Multidimensional Chromatography Techniques and Applications; Cortes, H. J., Ed.; Chromatographic Science Series; Marcel Dekker: New York and Basel, 1990; Vol. 50, pp 74-144. (11) Huber, L.; Obbens, H. J . Chromatogr. 1983, 279, 167-172. (12) Gordon, B. M.; Uhrig, M. S.; Borgerding, M. F.; Chung, H. L.; Coleman, W. M., 111; Elder, J. F., Jr.; Giles, J. A.; Moore, D. S.; Rix, C. E.; White, E. L. J . Chromatogr. Sci. 1988, 26, 174-180. (13) Krock, K. A,: Wilkins, C. L. Anal. rhim. Acta, in press.

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or selective qualitative and/or quantitative analysis of complex mixtures or for complete qualitative analysis of a complex mixture. This paper addresses the latter issue. Because of the time-consuming nature of MDGC, there have been a limited number of papers involving "total"qualitative analysis of a multicomponent mixture by sequential heart-cutting procedures. As an example, Gordon and co-workersanalyzed flue-cured tobacco essential oil by a sequential heart-cutting method and identified 306 compounds from among the total of 1500 detected. This analysis required 23 individual heart cuts and 48 h of instrument time.12 When such an intermediate trapping method is employed, it is necessary to perform n injections of the sample for a sequence of n heart cuts, with each injection followed by analysis of the heart cut in a secondary column. In this paper, it is proposed and experimentally demonstrated that parallel, rather than sequential, heart cuts are feasible, making it possible to significantly reduce the time required to carry out a multidimensional gas chromatographic separation with intermediate cryogenic traps. Before discussingthe concepts involved in a parallel cryogenic trapping system, it is worthwhile to describe the existing column switching systems utilized in MDGC. There are three alternative types of column switching techniques commonly employed in an MDGC system. The first is based upon use of multiport valves, the second uses pressure balancing to accomplishswitching between different columns, and the third employs a thermally modulated constant injection interface used in a way whichis significantly different from the first two techniques.14J5The fundamental differencebetween the valve and pressure balancing switching techniques is that, in the former, valves come into contact with the effluent, and when pressure switching is used, there are no valves and, consequently, no such contact occurs. The third method employsa valveless high-speedmodulator which achieves separation by thermally modulating a small section of a low-speed precolumn directly coupled to a high-speed column with different separation characteristics. As demonstrations of this method, a known test mixture and an unknown coal liquid sample were analyzed. Mixture component identification is based exclusively upon use of twodimensionalretention times and, as the investigators observed, has the potential for ambiguity inherent in basing identification upon such relatively nonspecific information. In its present form, this particular two-dimensional approach is incompatible with spectroscopic detection. Previously, on-line valves typically had large dead volumes, compromising chromatographic resolution and facilitating temperature-dependent adsorption of the sample on the valve walls. These problems lead to peak broadening and loss of peak intensity. With improvements in on-line valve technology, such difficulties are reduced, if not completely eliminated. It is recognized in the present investigation that a valve-based system could lead to significant sample adsorption a t high temperatures, resulting in partial or, in some cases, complete loss of one or more components. However, it is hoped that, in the future, commercialvalve manufacturers would incorporate design changes in light of the feasibility of the experimental strategy demonstrated here for analysis of complex mixtures by MDGC-IR-MS. On the other hand, in a recent paper Gordon, Rix, and Borgerding16compared three different commercially available column switchingdevices. Two of the three switching devices employ Deans principle of valveless switching (i.e., pressure 1986,57, 2779-2787, (15) Liu, 2.; Phillips, J. B. J. Chromtogr. Sci. 1991,29, 227-231. (16)Gordon, B. M.; Rix, C. E.: Borgerding, M. F. J. Chromatogr. Sci. 1985,23, 1-10.

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switching),and the third incorporates a four-port rotary valve. For systems based upon the Deans design," precise pressure balancing at the intermediate point is required, or if a restrictor is utilized, precise matching of the pressure drop across the restrictor and the analytical column is necessary. In contrast, the four-port rotary valve does not require precision controls and is simple to use. The only minor problem Gordon and co-workers reported was a slight shift in retention times of the peaks eluting after the heart cut.16 With respect to analyte adsorption, each system showed activity toward a t least one compound. It was concluded that the performance of the three systems was equivalent, at least for qualitative analysis purposes, and that multidimensional GC analysis can be carried out using a valve system without serious loss of chromatographic resolution. Parallel Cryogenic Trapping. As mentioned above, the technique of heart cutting by use of an intermediate cryogenic trap normally limits the number of cuts to one for a given injection. A method to increase the number of cuts per injection is to arrange a number of cryogenictraps in parallel, using a valve to feed a selected trap with analyte eluting from the primary column. In operation, this would be analogous to the technique employed in a railway switching yard, where one shunts newly arriving trains onto tracks that are empty. Experimental realization of this approach would be of significant benefit for complex mixture analysis by combined multidimensional gas chromatography, infrared, and mass spectrometry (MDGC-IR-MS). It would provide increased resolution of the cuts and simultaneously reduce the mass dynamic range and time required to carry out a complete qualitative analysis. Even though it is conceptually possible to have a large number of cryogenic traps in parallel, practically, the number of traps that can be arranged in parallel will be limited. However, a t least half a dozen (n = 6) such traps would present no difficult design problems. Analysis of the heart-cut material is accomplished by reinjection of heart cuts onto the analytical column by warming a single trap and then observing the chromatogram. For the multitrap system visualized, the procedure would be repeated until all the traps had been warmed. Furthermore, it would be possible to include another set of m traps parallel to the n traps. This arrangement could allow one to carry out heart cuts of a heart cut, and so forth. The use of computercontrolled valves and suitable computer algorithms would extend the versatility of such a system. Use of the separatory power of a MDGC chromatograph coupled with multispectral detection and modern computers could greatly enhance the reliability of qualitative analysis of complex unknown mixtures by GC-IR-MS. In this paper, experiments utilizing two parallel traps interposed between two GC columns are described.

EXPERIMENTAL SECTION Apparatus. The multidimensional gas chromatography system used in the present study is based upon a commercially available Hewlett-Packard (HP) GC-FTIR-MS system. This system includes an HP 5890 Series I1gaschromatograph,modified to incorporate dual parallel cryogenictraps; an HP 5965BFourier transform infrared detector (IRD);and a HP 5970Bma88 selective detector (MSD). The IRD and MSD are directly connected by a 1 : l O splitter with one part transferred to the MSD and 10parts to the IRD. Data are collected with two HP 300series computers running the HP IRD and MSD ChemStation software. A diagram of the plumbing for the multidimensional system is shown in Figure 1. The cryogenic traps are external to the gas chromatographic oven, and the external trap oven air temperature is maintained at approximately 200 "C thoughout trapping and reinjection. The cryogenic traps are fabricated from 1/16-in. (17) Deans, D.

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Vent

L Helium

Flgure 1. This is a diagram of the general plumbing for the dual parallel cryogenictrapping system used in this study. Not shown in the diagram are the trap extensions which allowed the liquid nitrogen to be vented out of the oven. The short ends of the traps were sealed with hightemperature adhesive to provide a unidirectional flow.

stainless steel tubing and are bent to provide a 10-cm trapping distance while shunting excess liquid nitrogen out of the oven. Both traps are placed in direct contact with the heater block, which is kept at a constant temperature of about 275 "C. Control of liquid nitrogen, trap flush gas, and analytical column makeup gas is accomplished with quick-acting toggle valves (Supelco, Bellefonte, PA). Supelco needle valves are used for fine control of the flush and makeup gases. In the MDGC system, both the precolumn and the separating column are placed serially inside the HP 5890 Series I1 GC (see Figure 1). A 30 m X 0.32 mm i.d. fused silica column coated with 14 % cyanopropylphenyl-86% methyl polysiloxane (DB-1701, 1.0 pm film thickness) (J & W Scientific, Folsom, CA) is used as the precolumn. The precolumn is connectedtoa (Valco,Houston, TX) three-port, two-way valve (300 "C maximum temperature) which can be switched between the detectors or the cryogenic liquid nitrogen traps. Trap selection is accomplished by use of high-performance onioff valves (SGE, Austin, TX), and the trapping is done in fused silica tubing with a DB-5 (5% phenylmethylpolysiloxane)stationary phase with a film thickness of 0.25 wm. The on/off valves are placed in front of the traps to eliminate any chromatographic peak broadening caused by sample contact with the valves or dead volume in the valves. Broadening caused by the valves is eliminated by condensingthe heart cut into a very narrow band in the trap. A flush flow of helium sweeps any leftover material from the cut into the appropriate trap to prevent cross-contamination between traps. All connections, after the traps, are made with glass Press-Tight connectors (Restek Corp., Bellefonte, PA) to ensure chromatographic integrity. Heart-cut samples are reinjected onto a 60 m X 0.32 mm i.d. DB-5 column with a film thickness of 0.25 pm (J & W Scientific, Folsom, CA). Ultrahigh purity helium (99.999% He) is used as the mobile phase with a head pressure of 80 kPa, providing a linear velocity of 33 cm/s for the initial separation. Analytical column makeup gas is set to a pressure of 140 kPa to provide a linear velocity of about 30 cm/s in the analytical column. Infrared spectra are obtained by using the HP IRD to collect 10 scans per spectrum with an optical resolution of 16 cm-I. This corresponds to the collection of one spectrum per second. The IRD has a 100-pL (1 mm i.d. X 12 cm) internally gold-coated Pyrex lightpipe that is maintained at 200 O C . The detector used in the IRD is a narrow band MCT detector with a spectral cutoff of 750 cm-I. The optics of the IRD are purged with dried nitrogen. The HP 5970B MSD is run in the full spectrum mode with analysis between 40 and 300 Da. Pressures in the MSD were approximately 1.5 X 1W Torr throughout all experiments. Sample. The sample analyzed in this study is an unleaded gasoline sample from an unknown source. The sample injections were 0.25-pL splitless injections with a 50 mL/min purge at 55 s into the run.

Flgure 2. First 20.0 min of the total IRD response chromatogram of the unleaded gasoline sample. The total number of countable peaks in the entirechromatogramIs approximately 1 10. The areas underlined and labeled are the cuts that were selected for further MDQC analysls.

Table I. Chromatographic Oven Parameters for Precolumn and Heart-Cut Seaarationa initial final temp time rate temp time (OC) (min) ("C/min) ( O C ) (rnin) precolumn 35 5 7 240 10 cut A 35 isothermal cut B 35 isothermal cut c 35 isothermal cut D 35 2 7 70 6 Procedure. The unleaded gasoline sample was first analyzed using a single-stage GC separation by the precolumn to obtain the total response chromatogram shown in Figure 2. This chromatogram, which contained about 50 peaks eluting within the first 20 min, was required to determine the timing for heartcut analysis. Thus, sample analysis was a two-step process in which the first step was simply a separation through the preliminary column, and the second step, using a second portion of sample, was the trapping and heart-cut analysis step. The chromatographicoven temperature parameters used in the study are summarized in Table I. During an analysis, the separation can be monitored by both the IRD and MSD, and at appropriate times, the two-way valve is turned to route column flow to the cryogenictraps. In operation, one of the traps is opened to allow the cut to be trapped, and simultaneously the vent is opened to maintain the 33 cm/s carrier gas linear velocity. At the end of the heart cut, the two-way valve is turned to direct flow to the detectors, and a little helium is leaked in through the flush line to purge the transfer lines of leftover sample. After both traps are filled,the GC oven temperature is reduced to the temperature required for the secondary separation, and a flow is established through the trap of interest by flowing helium through the trap and analytical column using the flush gas line. The precolumn head pressure is dropped to 35 kPa to allow the analytical column flow to reach the detectors. After the chromatographic baseline stabilizes, the selected trap is warmed by turning off the liquid nitrogen flow to the trap. With the trap heater block at approximately 200 "C, with both traps cooled down, the trap warm-up time is about 1-2 s.

RESULTS AND DISCUSSION The objective of this study is to investigate the advantage of using parallel cryogenic traps for the GC-GC-IR-MS qualitative analysis of a complex multicomponent mixture. Thus, the focus is second-stage GC separation of two adjacent segments of a chromatogram, trapped under cryogenic conditions, to yield increased chromatographic resolution

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