Instrumentation Peter R. Griffiths Stephen L. Pentoney, Jr. Aldo Giorgetti Kenneth H. Shafer* Department of Chemistry University of California Riverside, Calif. 92521
The impetus for combining chromatographs with spectrometers arose because the identification capability of chromatography alone is generally insufficient for the identification and quantification of every component of a complex mixture. The limitations of chromatography arise largely from the ambiguity of retention data. Typically, identification by chromatographic means alone requires some prior knowledge of the probable chemical structure of the components to be identified and the availability of authentic reference standards. These standards must therefore be predesignated for a particular analysis. Mixtures such as extracts of environmental samples often contain a multitude of components with a wide range of chemical diversity, the origin of which is generally unknown. The components may be present at levels from major to ultratrace, with a rapidly increasing number of compounds being observed as the detection limit is reduced. In many cases the identity of the major components is known, but the identity of minor chromatographic peaks (several of which may never have been observed previously in chromatograms of samples with a similar origin) can be of critical interest. The need for potentially thousands of standards for a conventional chromatographic analysis of samples of this type, together with the limited availability of many of these standards, makes the time (and cost) per analysis prohibitive.
give even this much information. Only when these spectrometers are linked to a chromatograph, so that each component is separated from the others, can the full power of MS or PT-IR for mixture analysis be realized. This linkage of two complementary analytical instruments was dubbed hyphenation in a farsighted review article by the late Tomas Hirschfeld (7). By combining the separation power of a chromatograph and the identification power of a spectrometer, it often becomes possible to identify many of the components of a complex mixture. After the various components have been identified, it becomes a relatively easy (albeit somewhat time-consuming) matter to determine the concentration of each through the use of calibration standards.
The HYPHENATION of CHROMATOGRAPHY FT-IR
SPECTROMETRY
A more certain identification of each peak can be achieved through the use of sensitive spectrometers to obtain a unique signature of each eluate. Ideally the spectrum should be measured in real time (i.e., in a time that 0003-2700/86/A358-1349$01.50/0 © 1986 American Chemical Society
is less than the width of the peak) as each component elutes, so that each measurement can be performed without trapping the sample. Mass spectrometry (MS) is the most commonly applied technique for characterizing chromatographic peaks. MS certainly merits its preeminent position in view of its high sensitivity and fast scan speed and the uniqueness of the mass spectra of many compounds. Despite its strengths, MS has certain drawbacks. For example, structural isomers often are not differentiated by MS, and library searches can sometimes lead to erroneous identifications. Under these circumstances, a complementary technique is desirable, and Fourier transform infrared (FT-IR) spectrometry has emerged as the most generally useful alternative—not only for gas chromatography (GC) but also for high-performance liquid chromatography (HPLC) and, more recently, supercritical fluid chromatography (SFC). When applied to a complex sample without a separation step, neither MS nor FT-IR spectrometry will lead to the identification or quantification of even one component. At best FT-IR can provide a general description of the chemical class of the major components, and mass spectra of such samples are so complex that they cannot
The hyphenated techniques share an important characteristic in that identification is accomplished with unique spectroscopic information rather than retention time data. The purpose of the spectrometry is predominantly identification, even though quantification is often also possible, for example from peak heights or areas measured from total ion current or Gram-Schmidt reconstructed chromatograms. The hyphenated techniques consequently do not have the limitations stated previously for chromatography or spectrometry applied alone, and complex mixtures often can be analyzed in detail with little or no prior chemical information This paper is dedicated to the memory of Tomas Hirschfeld, who coined the term hyphenated techniques and pioneered many of the advances. * Current address: Analect Instruments, Division of Laser Precision, 17819 Gillette Ave., Irvine, Calif. 92714
ANALYTICAL CHEMISTRY, VOL. 58, NO. 13, NOVEMBER 1986 · 1349 A
about the sample. In this paper, the ways in which gas, liquid, and supercritical fluid chromatographs have been interfaced to F T - I R spectrometers will be described. The extent to which the performance of the chromatograph can be matched to that of the spectrometer through the interface determines the analytical capabilities of any hyphenated technique. Spectroscopic sensitivity and information content, chromatographic resolution and capacity, and the time of analysis are all interdependent. In some interfaces, chromatographic selectivity has been compromised to improve spectral sensitivity. For example, for several HPLC/ FT-IR measurements, a particular mobile phase was selected based on its spectral transmission rather than its chromatographic performance. In successful interfaces, the performance of neither the chromatograph nor the spectrometer should be compromised if their combination is to become of general utility. Thus, many of the more recent developments in HPLC/ FT-IR have centered on eliminating the mobile phase prior to the FT-IR measurement so that the interface can be applied to separations performed with any mobile phase. Until a few years ago, GC/FT-IR systems were only available commer-
cially as accessories to top-of-the-line high-resolution FT-IR spectrometers. It is now becoming generally accepted that high resolution and high sensitivity are largely independent. Indeed many low-cost FT-IR spectrometers can have equally high (and sometimes higher) signal-to-noise ratio (S/N) compared with high-resolution instruments operating at well under their maximum resolution. It is also noteworthy that several contemporary bench-top GC/FT-IR systems fit in an area smaller than the space once required for the spectrometer alone. User convenience has also improved, in part because of the implementation of compact and inexpensive microcomputers instead of the larger, yet no more powerful, minicomputers that were installed in many FT-IR spectrometers manufactured before about 1981. The software for the hyphenated techniques that is now available from most manufacturers allows data acquisition and manipulation to be performed either in a totally automated or user-interactive mode. Simultaneous display of the spectral and chromatographic data has become quite commonplace because of improvements in both computer software and hardware. Spectral searching, especially for GC/FT-IR data, has become very fast
and can give less ambiguous identifications than GC/MS data (2, 3). However, it should be recognized that unequivocal identification depends on the availability of good reference data. Three libraries of vapor-phase reference spectra are available for GC/FTIR. The first is the U.S. Environmental Protection Agency's (EPA) data base, containing 3300 entries. This is a widely used library, because it is in the public domain, but it has a high proportion of errors. The second, which is sold by Sadtler Research Laboratories (Philadelphia, Pa.), contains more than 10,000 spectra, and the third (the Aldrich vapor-phase library) contains the spectra of 5100 compounds. Several other data bases contain digital reference spectra of compounds in the condensed phase, but there is far less uniformity in condensed-phase spectral libraries than in their vaporphase analogues. For some collections, samples have been prepared as KBr disks; for others mineral oil mulls were used. Some libraries contain only hard-copy spectra, whereas other spectra can be purchased in digital form. Some digitized data bases contain high-quality spectra measured on FT-IR spectrometers, but others (in particular the large data base of digital condensed-phase spectra marketed by Sadtler Research) contain grating
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spectra that were subsequently digi tized from the hard copy. Computerized identification of com pounds from their IR spectra has been performed in several ways. The most common method involves comparison of the absorbance spectrum of the un known with a library of reference spectra after scaling the absorbance of the most intense peak in the sample and reference spectra to the same val ue. This approach has been very suc cessful for gas-phase spectra, but the level of success for condensed-phase spectra is reduced by the presence of sloping base lines. Condensed-phase spectra have been converted to their first derivatives to reduce the effect of base line slope before starting the search. Another way of improving search re sults is to compare the Fourier trans form (FT) of the spectrum of each un known with the FT of all spectra in the data base. Both the early points and late points in these arrays may be truncated to eliminate the effect of base line slope and high-frequency noise, respectively. Although the effi cacy of this approach has been shown (4), it has still to be implemented on any commercial GC/FT-IR system. The sensitivity and data-processing capabilities of contemporary FT-IR spectrometers are at a very advanced
level. Scan speeds are sufficiently high that several interferograms can be av eraged per second, allowing real-time monitoring of peaks eluting from all types of chromatographs. In the re maining sections of this paper, the present state of the art of GC/FT-IR, HPLC/FT-IR, and SFC/FT-IR inter faces is described. We hope that this treatment shows the common features of these interfaces and provides an in dication of the directions that the de velopment of these devices is taking. Capillary GC/FT-IR
The most important component of most GC/FT-IR interfaces is an inter nally gold-coated glass tube, or light pipe, which serves as a gas cell through which the effluent from a capillary GC column flows. Spectra are measured continuously at inter vals of about 1 s, typically by signalaveraging blocks of 3-10 interfero grams per spectral data file. The di mensions of most light pipes designed for capillary GC/FT-IR are fairly sim ilar as the cell volume should be ap proximately equal to the volume of carrier gas contained between the half-height points of the GC peaks to obtain spectra with the optimum S/N (5). The actual peak volume depends on the internal diameter (i.d.) of the GC column and the thickness of the
stationary phase, but it is usually be tween about 50 and 200 μ\^. A suitable compromise between length and i.d. of the light pipe must be made if maxi mum performance is to be attained. A consensus among most instrument de signers now appears to have been reached, and most light pipes for cap illary GC/FT-IR are constructed from glass tubes with an i.d. of 1 mm and a length between 10 and 20 cm. With GC/FT-IR interfaces based on light pipes, identifiable spectra can usually be measured from injected quantities of strongly absorbing samples in the 5-25-ng range (6-8). For weakly ab sorbing compounds, however, it may be necessary to inject as much as 100 ng to obtain an identifiable spec trum. Although it is possible that the minimum identifiable quantity (MIQ) of strongly absorbing samples being characterized using light-pipe-based GC/FT-IR interfaces could be re duced to about 400 pg in a completely optimized system (9), it is quite un likely that any further decrease can be achieved without changing the funda mental nature of the GC/FT-IR inter face. One way in which the MIQ has been reduced to subnanogram levels has in volved trapping each separated com ponent in some fashion. One of the better ways of trapping GC eluates is
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1986
Figure 1. Gas chromatogram of 1-μΙ. injection of a herbicide still bottoms extract Spectra of peaks a through d are shown in Figure 2
to add a small quantity of argon to the column effluent and to pass the result ing mixture onto the surface of a met al substrate that has been cooled to about 12 K. Each solute is trapped in an argon matrix as the helium is pumped off. By slowly moving the sur face on which the matrix is being formed, a record of the chromatogram is made. After deposition, the sub strate is moved into the beam of an FT-IR spectrometer for the spec tral measurement of each GC peak (10-13). A commercial GC/FT-IR in terface based on this principle, known as the Cryolect, has resulted in com pounds with average absorptivities be ing identified for quantities in the range of 100-200 pg. For strong IR ab sorbers, even lower MIQs (10-50 pg) can be achieved. One of the keys to the high sensitiv ity of the matrix isolation approach is the very small area over which the sample is deposited. The effective thickness of a given amount of a GC component (or any other sample) is inversely proportional to its cross-sec tional area. With the Cryolect, each solute can be deposited in a spot whose diameter can be as small as 0.25 mm. Thus, all other factors being equal, the intensity of the bands due to a given quantity of a matrix-isolat ed GC solute trapped as a 0.25-mm-diameter spot should be 16 times greater than that of bands due to the same amount of the solute held in a 1-mm i.d. light pipe. The low temperature of the matrix also results in reduced widths and in creased absorbances of the spectral bands, which leads to a further de
crease in MIQ and an increase in specificity, relative to the light pipe interface. Because the point at which the solutes are deposited is separated by several centimeters from the focus of the IR beam, spectral data may not be acquired in real time. Although several hours may be required for a single GC/FT-IR run made with the matrix isolation interface, each com ponent is retained on the surface of the substrate for an indefinite period of time. The S/N of the spectra of mi nor GC peaks can therefore be in creased by extensive signal averaging. Three factors (low matrix tempera ture, small-area deposition, and trap ping to allow signal averaging) there fore contribute to the high sensitivity of the matrix isolation GC/FT-IR in terface. Of these, we will see that the last two are also of importance in sev eral other interfaces to be described. The subnanogram detection limits of the matrix isolation approach are not attained cheaply, and the substan tially higher cost of the Cryolect com pared with a light pipe interface has undoubtedly limited its use. In an at tempt to reap many of the benefits of matrix isolation without the cost, we have studied the feasibility of trap ping GC peaks at temperatures that are only a few degrees below ambient. In our approach the column effluent is passed onto a moving thermoelectrically cooled ZnSe window, which is held at a temperature between —10 and —45 °C. Each solute can be de posited as a spot of approximately 150-μπι diameter {14). The window is then moved into the focus of an F T IR microscope, so that the beam diam
1354 A · ANALYTICAL CHEMISTRY, VOL. 58, NO. 13, NOVEMBER 1986
eter can be matched to the size of the spot. Spectra are then measured a short time after deposition. MIQs for spectra measured with this interface are only slightly greater than those re ported for the matrix isolation inter face (15), whereas the cost is compara ble to that of the light pipe interface. At this point, the biggest disadvantage of the microscope-based interface is its inability to trap subnanogram quantities of volatile compounds be cause the lowest temperature to which the window has been cooled is —45 °C. To circumvent this problem, the inter face in our laboratory is being modi fied for operation at 77 K. The quality of the data that can be obtained with this interface can be il lustrated by spectra obtained from the chromatogram of the chloroform ex tract of the still bottoms from a herbi cide manufacturing plant. The sample was injected onto a 60-m-long, 0.25-mm-i.d. wall-coated open-tubular fused-silica column with a 0.25-/tmthick layer of DB-5, an intermediate polarity stationary phase. The chro matogram is shown in Figure 1, and the spectra of the four peaks indicated are shown in Figure 2. Although all these peaks have not yet been un equivocally identified, it is apparent that good spectra can be obtained from many of the separated compo nents. Because of the complementary na ture of IR and mass spectral data, more GC peaks can be identified when GC/FT-IR and GC/MS are used to gether than when either technique is applied separately (16-20). The devel opment of capillary GC/FT-IR inter-
Figure 2. IR spectra of four compo nents of the still bottoms extract shown as peaks a-d in Figure 1; note the com plete resolution of peaks b and c
faces that has taken place over the past decade has permitted detection limits to be reduced to the point at which IR spectra can now be obtained on many components of complex mix tures for which only mass spectra were obtainable in the past. Perhaps we are approaching the type of "black box" instrument in which a sample could be injected into a gas chromatograph and the identity and quantity of each sep arated component would be printed out shortly afterward. HPLC/FT-IR
To a large degree, the history of GC/ FT-IR development is mirrored in the development of HPLC/FT-IR inter faces. All early HPLC/FT-IR inter faces involved passing the column ef-
fluent through a flow cell and continu ously measuring spectra (21-24). In this respect, HPLC/FT-IR interfaces with this design can be considered to be analogous to light pipe GC/FT-IR interfaces. The key difference between GC/FT-IR and HPLC/FT-IR is that, whereas the mobile phase in GC is transparent to IR radiation, all HPLC mobile phases absorb IR radiation at certain wavelengths and some, such as water, have intense absorption across most of the spectrum. Absorption of the incident radiation by the solvent leads to several prob lems for the HPLC/FT-IR interface. First, solvent absorption bands must always be compensated if identifiable solute bands are to be observed. This effect completely obscures solute bands in several spectral regions and reduces the S/N at any wavelength where absorption by the solvent is ap preciable. Spectral subtraction of the solvent bands is almost impossible when the composition of the mobile phase is changing (e.g., during gradi ent elution). Finally, the maximum path length of the cell is determined by the ab sorption spectrum of the solvent. It is rare that the cell path length exceeds 200 μΐη without much of the useful IR spectrum being obscured by the sol vent absorption bands. Because the diameter of the flow cell should not exceed 4 mm, the largest cell volume is about 2.5 jtL, which is only about 1% of the width of a typical peak eluting from a 4.6-mm-i.d. column. For any interface between a chromatograph and an FT-IR spectrometer, the greatest sensitivity is found when the cell volume equals the full width at half height of the peak (5). Thus the limitation on path length imposed by the mobile phase places a significant limitation on the sensitivity of flow cell HPLC/FT-IR interfaces. For reversed-phase (RP) HPLC, where the mobile phase always contains water as a major constituent, the largest path length is about 25 μτη. In this case, the cell volume is reduced to 0.3 μL, and the sensitivity of the measurement is degraded proportionately. Thus, few useful R P - H P L C / F T - I R measure ments have been reported using a flow cell interface. One way of making the volume of flow cells and the width of HPLC peaks more compatible is to reduce the width of the peaks through the use of microbore columns. Taylor's group has reported some impressive HPLC/ FT-IR spectra obtained using such columns (25-28). The use of microbore columns does not get around the limitations imposed by the mobile phase completely, however, and most of the data reported by Taylor's group have necessitated the use of CDCI3 as
1356 A · ANALYTICAL CHEMISTRY, VOL. 58, NO. 13, NOVEMBER 1986
the mobile phase. It may be noted that the low solvent consumption of microbore columns means that expensive deuterated solvents can be used for microbore HPLC (AIHPLC) without solvent costs getting completely out of hand. Thus, for samples that can be separated using chloroform or other chlorinated solvents as the mobile phase, the application of microbore columns seems like a good solution to many of the problems of H P L C / F T IR. Jinno et al. (29) have even report ed using D2O as the mobile phase for /iHPLC/FT-IR measurements. Despite the promise of these ap proaches, flow cells appear to impose too many limitations on the HPLC/ FT-IR interface, and a much better approach is to eliminate the solvent prior to the IR measurement. Several approaches to solvent elimination have been described. They can be sep arated into two categories, involving either conventional or microbore HPLC columns. The first category necessitates elim inating the mobile phase emerging from 4.6-mm-i.d. columns, where the flow rate is typically on the order of 1 mL/min. A two-stage process has proved to be a useful way of eliminat ing organic solvents at this flow rate. In the first stage, 80-90% of the sol vent is evaporated, usually after nebulization of the column effluent. In the second stage, each solute is deposited onto a suitable substrate for measure ment of its IR spectrum. During depo sition, the remaining mobile phase must be completely evaporated to ob tain a spectrum free of solvent absorp tion bands. The higher the surface area of the substrate, the more rapidly the solvent can be eliminated. The use of powdered substrates has been fa vored because the transfer of heat through drops on a hot flat surface can be quite slow. After deposition, the diffuse reflectance (DR) IR spectrum of each solute is measured (30-32). Detection limits achieved using an interface based on this principle are about 2 orders of magnitude lower than can be obtained with a flow cell interface (33). Additionally all of the spectrum can be observed—not just those bands in the window regions of the solvent spectrum. This interface is not without its drawbacks, however. First, it is necessary to dispose of a considerable quantity of solvent va por. Second, it was noted earlier that the MIQ obtainable with a GC/FT-IR interface is strongly dependent on the area in which the sample can be de posited. This is equally true for an HPLC/FT-IR interface. When the so lution comes into contact with a pow dered substrate, it can spread rapidly by capillarity. To minimize the extent of spreading, it is usually necessary to
contain the powder in cups, and for the study of multicomponent mixtures, devices with many cups are needed. These have either been held in carousels (30-32) or linear transport devices (34, 35). One of these devices held 160 cups (31), and refilling even a small number of these cups after a chromatographic run proved to be an excessively time-consuming experience. The most common powders used as substrates for solvent elimination HPLC/FT-IR have been the alkali halides. Most alkali halides are very hygroscopic and can pick up water from the atmosphere while the solvent is being evaporated. Worse yet, they cannot be used at all if water is a component of the mobile phase, and a considerable amount of effort has been expended in the development of an HPLC/FT-IR interface suitable for RP-HPLC. Several approaches have been investigated for R P - H P L C / F T IR interfaces, based on the principle of solvent elimination. The simplest and most obvious is direct deposition on a substrate that is insoluble in water. Several candidates have been suggested, of which the most promising are diamond powder (industrial grade) and sulfur (36). If alkali halides are used, water must be completely eliminated before the elu-
ate comes into contact with the substrate. One approach involves adding 2,2'-dimethoxypropane at the end of the column, to remove water by the reaction: CH 3 C(OCH 3 ) 2 CH 3 + H 2 0 = CH 3 COCH 3 + 2 CH 3 OH The products of the reaction, acetone and methanol, can then be readily removed in the same fashion as normalphase solvents (34). In another approach, dichloromethane is added to the column effluent. The distribution ratio generally favors partition into the organic phase. Even if this is not the case, it is often possible to adjust the composition of the column effluent so that partition in the organic phase becomes favored (e.g., by addition of an acid or base). After separation and removal of the aqueous phase in the manner originally proposed by Karger et al. (37), CH2CI2 can be eliminated by evaporation and the DR spectrum of the solutes measured (38). Finally, it is also possible to separate water by means of a hydrophobic· membrane (39). None of these.solvent elimination approaches has proved popular enough to catalyze the development of a commercial HPLC/FT-IR interface. Although the detection limits are
quite acceptable, the amount of time needed to fill cups is not. We believe that an alternative solvent elimination device based on the use of microbore columns will prove more acceptable in the long term, as it should prove feasible to use an easily cleaned IR window or a metal plate as the substrate. The first solvent elimination interface between a microbore liquid chromatograph and an FT-IR spectrometer was the buffer memory technique of Jinno et al. (40-42). These workers deposited eluates from 0.3- and 0.5mm-i.d. packed fused-silica columns on a moving IR-transparent plate. Evaporation of the solvent was assisted by flowing nitrogen. The diameters of typical spots were approximately 2 mm, so that transmission spectra of the deposited solutes could be measured using a 4X beam condenser. Detection limits were about 100 ng per component. Jinno subsequently showed that it was possible to deposit solutes from RP-juHPLC columns onto metal mesh screens and to measure their spectra in an analogous fashion (43). Bearing in mind the criteria for reducing the MIQ for GC/ FT-IR components given in the previous section, it would appear that the best way to reduce detection limits in juHPLC/FT-IR is to decrease the diameter of the deposit below 0.5 mm. (continued on p. 1360 A) Performance
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Several methods of achieving this are being investigated in our laboratory, but none has yet allowed us to reach our goal of subnanogram detection limits.
SFC/FT-IR When a substance is raised above its critical temperature and pressure, a supercritical fluid is formed. The solvent power of a supercritical fluid is a function not only of its polarity but also its density. In turn, the density is affected by changes in temperature and pressure. SFC is unique among chromatographic techniques in that temperature, pressure, and composition programming can all be accomplished. The physical properties of supercritical fluids are intermediate between those of gases and liquids: Densities are liquidlike, viscosities are gaslike, and solute diffusivities are about 10 times greater in supercritical fluids than in liquids. If SFC is performed on packed columns, these properties enable higher linear velocities to be used in comparison with the corresponding HPLC separation, without loss in separation efficiency. More important, however, these properties allow wall-coated open-tubular columns to be used for SFC. The use of these columns has led to the current surge in popularity of SFC, because of
the increase in chromatographic resolution that can be achieved in comparison with packed columns (44). Like GC/FT-IR and HPLC/FT-IR, both flow cell and mobile-phase elimination approaches have been described for SFC/FT-IR. The use of flow cells for SFC/FT-IR is easier in some respects and more difficult in others than for HPLC/FT-IR. On the negative side, cells must be able to withstand pressures well in excess of 4000 psi. Few IR spectra of supercritical fluids have been reported in the past, so that the IR transparency of all mobile phases potentially applicable to SFC/FT-IR has not been well characterized. On a more positive note, by far the most popular fluid used to date for SFC has been CO2, and the transmittance of supercritical CO2 is significantly greater than that of most HPLC mobile phases, allowing cells of increased path length to be used. Thus flow cell SFC/FT-IR measurements would be expected to have greater sensitivity and yield more spectral information than the corresponding HPLC/FT-IR measurements, and several reports of flow cell S F C / F T IR measurements with CO2 as the mobile phase have appeared over the past four years (45, 46). The absorptivities of bands in the spectrum of supercritical C 0 2 have
been observed to increase with pressure, with the intensities of some symmetry-forbidden bands being more sensitive to pressure than the intense fundamentals (where the spectrum is "blacked out" in any event). Even though the problems of solvent compensation encountered in pressureprogrammed SFC/FT-IR are not as severe as those with gradient elution HPLC/FT-IR, a single reference spectrum still cannot be used throughout the analysis to compensate for absorption by the mobile phase, so that most SFC/FT-IR measurements using flow cells have been performed isobarically. French and Novotny (47) have proposed substituting supercritical xenon for CO2 to circumvent the problems imposed by the absorption bands of CO2. The chromatographic properties of C 0 2 and xenon are surprisingly similar, and preliminary results are promising. However, CO2 and xenon are both nonpolar and cannot be used for the separation of strongly polar analytes. In this case, polar mobile phases such as the Fréons, NH 3 , or SO2 will probably have to be used. Because these compounds are strong IR absorbers, the flow cell approach for SFC/FT-IR will have the same limitations that it does for HPLC/FT-IR. Johnson et al. (48) have suggested performing separations with two different
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1986
supercritical fluids (CO2 and Freon23), chosen on the basis of their IR spectra, so that IR opaque regions in the spectrum of one fluid are windows in the spectrum of the other. The dis advantages of this approach are that not only must two chromatograms be run, but the eluent strength of the two mobile phases must be very similar. Although CO2 and Freon-23 have sur prisingly similar chromatographic properties, this will not generally be the case, and the assignment of corre sponding peaks in the two chromato grams will probably prove trouble some. Although flow cell interfaces have several disadvantages with regard to the analysis of complex samples, their use in selected applications, such as on-line process monitoring, can be beneficial (49). In this type of applica tion, a few compounds of known struc tures can be monitored by selecting a mobile phase that not only affords an adequate separation but that also has the appropriate spectral windows. For complex sample analysis, mo bile-phase elimination appears to have several advantages for SFC/FT-IR. In
the first such interface for SFC/FTIR, the eluate was deposited on KC1 powder in a fashion similar to the ear lier HPLC/FT-IR interfaces, and DR spectra of the components were mea sured (50, 51). For this work, separa tions were performed on a 1-mm-i.d. column packed with ΙΟ-μτα. silica using CO2 with 5% CH3OH as the mobile phase. A great advantage of this ap proach for SFC/FT-IR results from the volatility of the mobile phase on releasing the pressure at the restrictor. It was observed that each component could be readily deposited in a spot of approximately 1 mm diameter if the substrate remained stationary. Alter natively, solutes could be deposited continuously on a moving substrate, thereby obviating the need for dis crete sample cups. An investigation of the MIQ that can be expected from the approach showed that identifiable spectra could be obtained from ap proximately 50 ng of each component of a mixture of quinones. As a rule of thumb, the critical tem perature is approximately equal to 1.5 times the boiling temperature in Kel vins. To keep the temperature of the
column reasonably low, most mobile phases for SFC are gaseous at stan dard temperature and pressure. Thus, on releasing the pressure at the re strictor, the mobile phase evaporates immediately. With the slow flow rates involved in capillary SFC with nar row· bore columns, the rapid evapora tion of the mobile phase allows solutes to be deposited as spots of about 250 μπι diameter when the column ef fluent passes through a narrow-bore restrictor. As a result, SFC/FT-IR measurements have been performed under a microscope in the same man ner described above for GC/FT-IR (52, 53) or using a technique similar to the buffer memory method for HPLC/ FT-IR(54). To illustrate the quality of SFC/ FT-IR spectra measured in this man ner, a capillary supercritical fluid chromatogram of the same sample whose gas chromatogram was illus trated in Figure 1 is shown in Figure 3.
Figure 3. Supercritical fluid chromatogram of a 0.06-μΙ. injection of the same sam ple shown in Figure 1 Column: 20 m long, ΙΟΟ-μΓη-i.d. fused silica, with 0.40-μπι layer of DB-5; oven temperature: 120 °C; pressure program: indicated on abscissa. Spectra of peaks a-d are shown in Figure 4
1362 A · ANALYTICAL CHEMISTRY, VOL. 58, NO. 13, NOVEMBER 1986
Figure 4. IR spectra of components a-d in Figure 3
Formaldehyde Rewnec from Wood Products
S F C / F T - I R s p e c t r a of four of t h e c o m p o n e n t s a r e s h o w n in F i g u r e 4, including t h a t of o n e c o m p o n e n t p r e s e n t as t h e leading edge of a n unresolved d o u b l e t (Figure 4b). M o s t of t h e p e a k s in t h e supercritical fluid c h r o m a t o g r a m do n o t c o r r e s p o n d t o p e a k s in t h e gas c h r o m a t o g r a m . T h e early-eluting p e a k a c h a r a c t e r i z e d b y S F C / F T IR, however, h a s t h e identical spect r u m as t h e late-eluting p e a k d in t h e G C / F T - I R d a t a , so it can be d e d u c e d t h a t m o s t p e a k s in t h e S F C c h r o m a t o gram are not attributable to the same c o m p o n e n t s giving rise t o p e a k s in t h e gas c h r o m a t o g r a m .
Conclusion
B. Meyer, Editor University of Washington B.A. Kottes Andrews and R.M. Reinhardt, Editors U.S. Department of Agriculture Summarizes current information on problems relating to measuring, abating, and understanding formaldehyde emissions from wood products bonded with formaldehydebased adhesive resins. Discusses the success of emission-reduction efforts, explores the problems remaining to be solved, and examines improvements in the understanding of mechanisms of formaldehyde generation and release. A useful resource for textile chemists and manufacturers, wood chemists and wood product manufacturers, industrial hygiéniste, architects, construction engineers, consumers, public interest groups, and regulatory groups. CONTENTS Formaldehyde Release: An Overview · Plywood and Wood-Based Panel Products · Wood-Panel Products Bonded with Phenol Formaldehyde Adhesives · Release Rate Coefficients from Selected Consumer Products · Cellulose Reaction with Formaldehyde * Cellulose Models for Formaldehyde Storage in Wood · Urea-Formaldehyde Resins · Mechanisms of Release from Bonded Wood Products · Automated Flow Injection Analysis System · Enzymatic Methods for Determining Formaldehyde Release · Formaldehyde Release from Building Materials · Large-Scale Test Chamber Methodology · Predicting Real-Life Formaldehyde Release · Tannin-Induced Formaldehyde Release Depression · Effect of Diffusion Barriers on Formaldehyde Emissions · European Formaldehyde Regulations · Occupational and Indoor Air Formaldehyde Exposure Based on a symposium sponsored by the Division of Cellulose, Paper, and Textile Chemistry of the American Chemical Society ACS Symposium Series No. 316 240 pages (1986) Clothbound LC 86-14194 ISBN 0-8412-0982-0 US & Canada $49.95 Export $59.95 Order From: American Chemical Society Distribution Office Dept. 24 1155 Sixteenth Street, N. W. Washington, DC 20036 or CALL TOLL FREE 800-424-6747 and use your credit cardl
It is obvious from t h e p r e c e d i n g discussion t h a t m a n y significant developm e n t s have t a k e n place recently in GC/FT-IR, H P L C / F T - I R , and S F C / F T - I R . It s h o u l d b e equally a p p a r e n t t h a t b y n o m e a n s all t h e p r o b l e m s involved in t h e design of a n interface s u i t a b l e for all t y p e s of c h r o m a t o g r a p h y have been solved. P r a c t i c a l evid e n c e for t h i s s t a t e m e n t can be found in t h e lack of a solvent e l i m i n a t i o n interface for H P L C / F T - I R a n d S F C / F T - I R from a n y c o m m e r c i a l source. Good designs for such interfaces a r e j u s t s t a r t i n g t o be r e p o r t e d , a n d we believe t h a t within a year or two t h e s e devices will begin t o a p p e a r in t h e marketplace. The recent developments in GC/FT-IR and SFC/ FT-IR taking place in the authors' laboratory were supported in part by Cooperative Agreements CR811730-02 and CR812258-01 between the University of California, Riverside, and the U.S. EPA's Environmental Monitoring Systems Laboratory, Las Vegas, and by a grant from the University of California Toxic Substances Program. This paper has been reviewed in accordance with the U.S. EPA's peer and administrative review policies and approved for presentation and publication. Mention of trade names does not constitute endorsement or recommendation for use. S.L.P. gratefully acknowledges support by an ACS Summer Analytical Fellowship, sponsored by Calgon Corporation.
References (1) Hirschfeld, T. Anal. Chem. 1980, 52, 297-312 A (2) Shafer, K. H.; Hayes, T. L.; Brasch, J. W.; Jakobsen, R. J. Anal. Chem. 1984, 56, 237-40. (3) Gurka, D. F.; Hiatt, M.; Titus, R. Anal. Chem. 1984,56,1102-10. (4) de Haseth, J. Α.; Azarraga, L. V. Anal. Chem. 1981,53, 2292-96. (5) Griffiths, P. R. Appl. Spectrosc. 1977, 31, 284-88. (6) Azarraga, L. V.; Potter, C. A. J. High Res. Chromatogr. Chromatogr. Coram. 1981,4,60-69. (7) Kuehl, D.; Kemeny, G. J.; Griffiths, P. R. Appl. Spectrosc. 1980, 34, 222-24. (8) Griffiths, P. R.; de Haseth, J. Α.; Azar raga, L. V. Anal. Chem. 1983, 55,136187 A. (9) Hirschfeld, T. Appl. Spectrosc. 1985, 39, 1086-87. (10) Mamantov, G.; Wehry, E. L.; Kemmerer, R. R.; Hinton, E. R. Anal. Chem. 1977, 49, 86-90. (11) Reedy, G. T.; Bourne, S.; Cunning
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ham, P. T. Anal. Chem. 1979,51,153540. (12) Schneider, J. F.; Reedy, G. T.; Ettinger, D. G. J. Chromatogr. Set. 1985, 23,49-53. (13) Reedy, G. T.; Ettinger, D. G.; Schneider, J. F. Anal. Chem. 1985,57, 1602-9. (14) Shafer, K. H.; Griffiths, P. R.; Fuoco, R. J. High Res. Chromatogr. Chroma togr. Comm. 1986,9,124-26. (15) Fuoco, R.; Shafer, Κ. Η.; Griffiths, P. R. Anal. Chem., in press. (16) Crawford, R. W.; Hirschfeld, T.; Sanford, R. H.; Wong, C. M. Anal. Chem. 1982,54,817-20. (17) Gurka, D. F.; Betowski, L. Anal. Chem. 1982,54,1819-24. (18) Wilkins, C. L.; Giss, G. N.; White, R. L.; Brissey, G. M ; Onyiriuka, E. C. Anal. Chem. 1982,54, 2260-64. (19) Laude, D. Α.; Brissey, G. M.; Ijames, C. F.; Brown, R. S.; Wilkins, C. L. Anal. Chem. 1984,56,1163-68. (20) Laude, D. Α.; Johlman, C. L.; Cooper, J. R.; Wilkins, C. L. Anal. Chem. 1985, 57,1044-^9. (21) Kizer, K. L.; Mantz, A. W.; Bonar, L. C. Am. Lab. 1975, 7(5), 85-97. (22) Vidrine, D. W.; Mattson, D. R. Appl. Spectrosc. 1978,32, 502-6. (23) Vidrine, D. W. J. Chromatogr. Sci. 1979,77,477-82. (24) Brown, R. S.; Taylor, L. T. Anal. Chem. 1983,55, 723-30. (25) Brown, R. S.; Taylor, L. T. Anal. Chem. 1983,55,1492-97. (26) Johnson, C. C ; Taylor, L. T. Anal. Chem. 1983,55,436-41. (27) Amateis, P. G.; Taylor, L. T. Anal. Chem. 1984,56,966-71. (28) Johnson, C.C.; Taylor, L. T. Anal. Chem. 1984,56, 2642-47. (29) Jinno, K.; Fujimoto, C ; Vematsu, G. Am. Lab. 1984, 76(2), 39-45. (30) Kuehl, D.; Griffiths, P. R. J. Chroma togr. Sci. 1979,17,471-76. (31) Kuehl, D.; Griffiths, P. R. Anal. Chem. 1980,52,1394-99. (32) Conroy, C. M.; Griffiths, P. R.; Jinno, K. Anal. Chem. 1985,57, 822-25. (33) Brown, R. H.; Knecht, J.; Witek, H. Proc. Soc. Photo-Opt. Instrum. Eng. 1981,289, 51-52. (34) Kalasinsky, K. S.; Smith, J.A.S.; Kalasinsky, V. F. Anal. Chem. 1985, 57, 1969-74. (35) Kalasinsky, V. F.; Smith, J.A.S.; Kal asinsky, K. S. Appl. Spectrosc. 1985, 39, 552-54. (36) Brackett, J. M.; Azarraga, L. V.; Cas tles, Μ. Α.; Rogers, L. B. Anal. Chem. 1984,56,2007-10. (37) Karger, B. L.; Kirby, D. P.; Vouros, P.; Foltz, R.; Hidy, B. Anal. Chem. 1984, 56, 2636-42. (38) Conroy, C. M.; Griffiths, P. R.; Duff, P. J.; Azarraga, L. V. Anal. Chem. 1984, 56, 2636-42. (39) Johnson, C. C ; Hellgeth, J. W.; Tay lor, L. T. Anal. Chem. 1985,57,610-15. (40) Jinno, K.; Fujimoto, C. J. High Res. Chromatogr. Chromatogr. Comm. 1981, 4, 532-33. (41) Jinno, K.; Fujimoto, C ; Hirata, Y. Appl. Spectrosc. 1982,36, 67-69. (42) Jinno, K.; Fujimoto, C ; Ishii, D. J. Chromatogr. 1982,239, 625-33. (43) Fujimoto, C ; Oosuka, T.; Jinno, K. Anal. Chim. Acta 1985,178,159-67. (44) Novotny, M.; Springston, S. R.; Peaden, P. Α.; Fjeldsted, J. C ; Lee, M. L. Anal. Chem. 1981,53, 407-14 A. (45) Shafer, K. H.; Griffiths, P. R. Anal. Chem. 1983, 55, 1939-42. (46) Olesik, S. V.; French, S. B.; Novotny, M. Chromatographia 1984,18, 489-95. (47) French, S. B.; Novotny, M. Anal. Chem. 1986,58,164-66. (continued on p. 1366 A)
(48) Johnson, C. C; Jordan, J. W.; Taylor, L. T.; Vidrine, D. W. Chromatographia 1985,20,717-23. (49) Skelton, R. J.; Johnson, C. C; Taylor, L. T. Chromatographia 1986,21,3-8. (50) Shafer, K. H.; Pentoney, S. L.; Grif fiths, P. R. J. High Res. Chromatogr. Chromatogr. Comm. 1984, 7, 707-9. (51) Shafer, K. H.; Pentoney, S. L.; Grif fiths, P. R. Anal. Chem. 1986,58,58-64. (52) Pentoney, S. L.; Shafer, K. H.; Grif fiths, P. R. J. High Res. Chromatogr. Chromatogr. Comm. 1986,9,168-71. (53) Pentoney, S. L.; Shafer, K. H.; Grif fiths, P. R. J. Chromatogr. Sci. 1986,24, 230-35. (54) Fujimoto, C; Hirata, Y. H.; Jinno, K.
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Peter Griffiths (left) is professor of chemistry at the University of Cali fornia, Riverside (UCR). His research interests include the application of FT-IR spectrometry to environmen tal, surface, and coal chemistry, the theory and practice of diffuse-reflec tance spectrometry in the mid- and near-infrared, and the development ofSFC. Stephen Pentoney (left center) re ceived his B.S. from California State University, Long Beach, and is now a fourth-year graduate student at UCR, where he is working on the de velopment of SFC and its analytical
applications. At UCR, he received a Sohio fellowship in analytical chem istry in 1983 and a Shell fellowship in 1984, and this summer he was award ed an ACS Division of Analytical Chemistry fellowship. Aldo Giorgetti (right center) received his Diploma of Chemistry and Ph.D. from the University of Lausanne, Switzerland, and is a postdoctoral as sociate at UCR. His research interests include the study of crown ether com plexes of rare earths and the develop ment of coupled techniques such as GC/FT-IR and the combination of
GC and SFC for the separation of mixtures of volatile and nonvolatile molecules. Kenneth Shafer (right) received his B.S. and M.S. degrees from Ohio Uni versity, after which he was employed at Battelle's Columbus Laboratories for seven years. He returned to Ohio University to obtain his Ph.D. and until recently was a postdoctoral as sociate at UCR. His main research in terest is the development of hyphen ated techniques. Currently he is man ager of FT-IR peripherals at Analect Instruments.
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