Anal. Chem. 1993, 65, 3290-3294
3290
Binary Mobile Phases for Supercritical Fluid Chromatography with Helium Microwave-Induced Plasma Detection Yubang Wang and Jon W . Camahan’ Department of Chemistry, Northern Illinois University, DeKalb, Illinois 60115
Microwave-induced plasma atomic emission detection is examined for supercritical fluid chromatography detection with binary methanol/ carbon dioxide mobile phases. Spectra are obtained and compared with pure helium plasmas, plasmas with added carbon dioxide, and added binary mobile phases. Trends of molecular emission intensities and species were examined as functions of methanol/carbon dioxide composition ratios and supercritical fluid pressures. Adding carbon dioxide to the pure helium plasma reduced and/or eliminated molecular bands from OH and N2+while producing intense bands of CN,C2,NH, and Nt. The addition of methanol to the carbon dioxide “doped” plasma produced intense OH emission while depressing the intensities of the other aforementioned molecular bands. Emission from OH increased significantly with additional methanol. As the pressure of the methanol/carbon dioxide mobile phase was increased, the general trend was that molecular band intensities tended to decrease. The effect was attributed to plasma cooling. The plasma easily tolerated the mixed mobile phases despite the added spectral complications of these molecular emissions. The advantages of the binary mobile phases for chromatography are illustrated with separations of moderately high molecular weight chlorinated organic compounds. Retention times are reduced and detectabilities are in the range of 1.4-1.9 ng/s for pchlorobiphenyl, 2,4‘-dichlorobenzophenoneand hexachlorobenzene. INTRODUCTION Supercritical fluid chromatography (SFC) has been recognized as an attractive and complementary method for compounds not amenable to separation and detection by gas chromatography (GC). With supercritical fluid mobile phases, large and relatively nonvolatile molecules can be solvated to provide efficient separations in much shorter analysis times than with liquid chromatography (LC). Recent advances in the development of SFC have stimulated research for compatible detection techniques. The ideal universal detector for SFC remains elusive. This detector should exhibit good analytical performance with various supercritical fluid mobile phases and chromatographic conditions. While nearly every GC or LC detector has been investigated for SFC, the most widely used SFC detectors are ultraviolet/visible molecular absorption and flame ionization detection (FID). Limitations of absorption detection include moderate sensitivity and limited selectivity. The FID is constrained to fluids yielding significant ionization during
* Author to whom correspondence should be addressed. 0003-2700/93/0365-3290S04.00/0
detection; polar supercritical fluid modifierssuch as methanol reduce or eliminate the capabilities of ionization-based detection. Element-selectivedetection based on atomic emissionoffers many potential advantages. Atomic emission lines are narrow and often intense, affording high selectivity and sensitivity. The most common chromatography detector based on these principles is the microwave-induced plasma (MIP). Application of the MIP as a GC detector was first reported by McCormack, Tong, and Cookel and Bache and Lisk.2 In the past several years, the MIP has proven to be particularly useful as an element-selective detector for GC.9-8 The MIP is relatively inexpensive, easily constructed, and compatible with gaseoussample introduction. The helium-sustainedMIP is particularly attractive because both nonmetals and metals can be detected at low concentration levels. These features make the MIP an attractive alternate detector for SFC. The f i i t examinationsof helium plasma atomic emissiondetection were initiated by Galante et al.7~8 Subsequently, studies have been performed in the laboratories of Lee? Long,lo and Carnahan.llJ2 The SFC mobile phases most commonly studied with atomic emission detection have been nonpolar fluids such as pure C02 and N2O. These nonpolar mobile phases constrain SFC applications to the separation of relatively nonpolar organic compounds. While other single-component supercritical fluid mobile phases such as ammonia or methanol offer polarity, difficulties involving supercritical-phase parameters and reactivity hamper the use of these potential mobile phases. With this in mind, polar fluid modifiers have been used to increase the solvent strength of low-polarity fluids such as COZ. The most commonly used modifier for C02 is methanol.13 The modified binary mobile phase alters chromatographic retention and selectivity, allowing more efficient and rapid separation for polar compounds while obtaining Gaussian peak shapes. Motley and Longlo examined the effects of methanol modifier with a carbon dioxide SFC mobile phase on the detection of iron as ferrocene. (1)McCormack, A. J.; Tong, S. C.; Cooke, W. D. Anal. Chem. 1966, 37,1470-1477. (2)Bache, C.A.; Lisk, 0. J. AMI. Chem. 1966,37,1477-1480. (3)Mohamad, A. H.; Caruso, J. A. In Adoances in Chromatography; Giddings, J. C., Gruehka, E., Brown, P. R., Eds.; Marcel Dekker: New York, 1987;Vol. 26,Chapter 5. (4)Element-Specific Chromatographic Detection by AtomicEmission Spectroscopy; Uden, P. C., Ed.; ACS Symposium Series 479;American Chemical Society: Washington, 1992. (5) Beenakker, C. I. M. Spectrochim. Acta, Part B 1976,31,483-486. (6)Quimby, B. D.;Sullivan, J. J. Anal. Chem. 1990,62,1027-1034. (7)Luffer, D. R.;Galante, L. J.;David, P. A,; Novotny, M.; Hieftje, G. M. Anal. Chem. 1988,60,1365-1369. (8)Galante, L. J.; Selby, M.; Luffer, D. R.;Hieftje, G. M.; Novotny, M. Anal. Chem. 1988,60,1369-1376. (9) Skelton, P. B.; Farmworth, P. B.; Markides, K. E.; Lee, M. L. Anal. Chem. 1989,61,1815-1821. (10)Motley, C. B.;Long, G. L. Appl. Spectrosc. 1990,44,667-672. (11)Zhang, L.; Carnahan, J. W.; Winans, R. E.; Neill, P.H. Anal. Chem. 1991,63,212-216. (12)Webster, G. K.;Camahan, J. W. Anal. Chem. 1992,64, 50-56. (13)Chester, D.L.; Pinkston, J. D.; Raynie, D. E. Anal. Chem. 1992, 64,153R-170R. 0 1993 American Chemlcal Soclety
ANALYTICAL CHEMISTRY, VOL. 65, NO. 22, NOVEMBER 15, lQS3
9291
steel tube union Flgure 1. SFC/MIP Interface and plasma torch. Helium
Improvements in signal-to-noise ratios were observed in the presence of the modifier. Detailed effects of modifiers upon plasma spectral and analyte excitation characteristics have not been examined. Studies of this nature are mandatory if applications to polar nonvolatile molecules are to be addressed. In this work, a moderate-power microwave-induced plasma is explored as a chlorine-selective detector for packed-column SFC with binary mobile phases. Background emission in the 200-600nm spectral region is investigated with pure helium discharges and those with C02 and Codmethanol phases. Major molecular bands and atomic emission lines are identified. Mobile-phase effects upon molecular band background emission and plasma stability are examined. The chlorine emission line at 479.5 nm is selected to monitor eluted chlorinecontaining compounds. The optimal plasma operating conditions are defined. Analytical characteristics of the detector are monitored with changes in chromatographic parameters such as SFC pressures and mobile-phase composition. EXPERIMENTAL SECTION The supercritical fluid chromatography/helium microwaveinduced plasma detection system consisted of four major components: a supercritical fluid chromatograph, a 500-W microwave-induced plasma, the SFC/MIP interface, and an optical detection system. With the exception of the optics, a description of the system may be found in ref 12. Selected details follow. SFC/MIP Interface and Plasma Torch. The plasma was sustained inside a 8.5 mm i.d. X 11.0 mm 0.d. quartz torch. The interface is shown in Figure 1. The White and Associates (Pittsburgh, PA) 150 X 1 mm Deltabond Phenyl 5-pm packed SFC column outlet was connected to a 7-10 mL/min integral restrictor from White and Associates or a frit restrictor (Lee Scientific, Inc.) of 1.2 cm/s (specifiedwith a column temperature of 75 O C and C02 mobile phase at 75 atm) flow rate. The restrictor was routed through a Swagelok union and a stainlesstube (0.25mm i.d.) centered in the Teflon insert of the torch. The Teflon insert fits snugly into the quartz tube, which is sleeved by a second Swagelok union and connected to the torch. Teflon tape is used to create the seal between the quartz tube and the nut. To avoid mobile-phase freezing due to Joule-Thomson cooling at the restrictor tip, the restrictor is placed close to the plasma. The minimum distance was maintained from 1to 3 cm to avoid restrictor damage. Additionally, the quartz portion of the torch was wrapped by resistively heated Nichrome wire. Spectrometer. A 0.34-m focal-length Model 340E monochromator (SpexIndustries,Edison, NJ) with a 1200groove/” grating (reciprocal linear dispersion of 2.5 nm/mm) was used. The photomultiplier was a Model R928 (Hamamatsu Corp., Toyooka Village, Japan) biased at -1100 V. Spectrometer entrance and exit slits were set at 50 pm. The detector output was converted to a voltage, filtered, and amplified by an operational amplifier based system built at NIU. Data collection was performed with a Fisher Series 500 strip chart recorder. SFCMobilePhaee. Another concemwithmixedmobilephases is that T,and P, are different from that of the one-component phase. Considerations must be given to predict T,and P, for solvent mixtures. Crowther and Henionl‘ have calculated the minimum temperature and pressure necessary for supercritical conditione for various mole fractions of methanol modifier. For (14) Crowther, J. B.;Henion, J. D. Anal. Chem. 1986,57,2711-2716.
200
400
600
Wavelength (nm) Flgure 2. Spectrum of pure hellum discharge.
10% methanol in C02, it can be estimated that T,is 57 O C and P, is 85 bar. This information is useful to define pump control parameters and operationaltemperatures. Two sources of binary mobile phases were used: 10% methanol in C02 was purchased from Scott SpecialtyGases; 6.5 and 19.2% (v/v)methanol in C02 were prepared in the syringe pump. Reagents. Pure helium (99.995%)was purchased from Great Lakes Airgas, Inc. (West Chicago, IL). Research grade 1,3dichlorobenzene,p-chlorobiphenyl, 2,4’-dichlorobenzophenone, and hexachlorobenzenewere obtained from Aldrich (Milwaukee, WI).
RESULTS AND DISCUSSION Spectrum of P u r e Helium Discharge. A spectrum of a pure helium plasma was obtained. The spectrum is similar to that obtained by Tanabe, Hariguchi, and Fuwa.16 As shown in the Figure 2, the major line emissions arise from atomic helium. Intense helium lines are seen at 388.9,402.6,447.1, 492.2, 501.6, and 587.6 nm. Other emission lines as well as a number of molecular emission bands are seen due to impurities in the helium gas and species from air entrained into the plasma region. The hydrogen atomic emission lines at 434.0 and 486.1 nm are strong. The OH band at 306.4 nm and the NH bands a t 336.0 and 337.0 nm are relatively strong. There ia a weak Nz band at 357.7 nm. Bands of N2+ appear a t 391.4 and 427.8 nm. A C2 band appears at 563.6 nm. The second-order 237.0-nm NO band head appears at 474.0 nm. In general, helium is a good support gas for plasma emission spectrometry as the emission background is relatively “clean”. Spectrum of Carbon Dioxide Doped Plasma. As seen in Figure 3, the addition of the carbon dioxide mobile phase has a dramatic effect on the spectral background. For these measurements, C02 was introduced through an SFC restrictor with a pump pressure of 250 atm. The helium plasma gas flow rate was 18 L/min, the forward power was 500 W, and the reflected power was 0 W. Helium and hydrogen line emission intensities were reduced to the point that they were not observed under these experimental conditions. The firstand second-order 247.8-nm carbon atom line emissions are intense. (The second-order emission appears at 495.7 nm.) Band emissions from OH and N2+ were reduced significantly and/or eliminated. Several intense molecular bands appear dueto CN (359.0,388.3, and 421.6nm1, C2 (436.5,473.7,512.9, 516.5,550.2,554.0 558.6, and 563.6 nm), NH (336.0 and 337.0 nm), and N2 (405.9 nm). Of these, CN and CZdominate the spectrum. These background features may preclude or interfere with the line emission measurement. Spectrum of Methanol/COz Binary Mobile Phase Doped Plasma. For these experiments, methanol was injected into the pump cylinder and an integral restrictor (16) Tanabe, K.;Hariguchi, H.; Fuwa, K. Spectrochim. Acta 1981, 36B, 119-127.
ANALYTICAL CHEMISTRY, VOL. 65, NO. 22, NOVEMBER 15, 1993
9292
U
I
200
I
I
400
600
Wavelength (nm) Flgure 8. Spectrum of pure C02introduced into the helium discharge. The integral restrlctor was used with a pressure of 250 atm.
I
I
I
200
400
600
Wavelength (nm) Flgure 1. Spectrum of 19.2% methand/80.8% C02 introduced Into the helium discharm The intecrral restrictor was used with a mssure of 250 atm.
-
-
tl
.-* c v)
C
Q)
c
C
I
200
I
I
I
400
600
200
Wavelength (nm) Figure 4. Spectrum of 6.5% methanoi/93.5% C02 introduced into the helium discharge. The integral restrictor was used with a pressure of 250 atm.
was used. The pump pressure was 250 atm. For the 6.5% methanol mixture, 10 mL of methanol was injected into the 180-mLsyringe. The remaining volume was filled with liquid COS. Figure 4 shows the spectrum of 6.5% methanol/93.5% C02 doped in the plasma. The addition of methanol yielded intense OH molecular emission at 306.0 nm. As with Figure 3, the spectrum is dominated by C2 and CN molecular emissions. However, the intensities of all CN bands are reduced. Interestingly, the intensities of the C2 bands at 473.7, 516.5, and 563.6 nm (2.62-, 2.40-, and 2.40-eV upper state energies, respectively) decreased while the C2 band at 436.5 nm (upper state energy, 3.32 eV) increased. The increased intensity of the higher energy transitions might be due to near-resonant energy coupling with the 4.05-eV upper state of the OH moiety. Figure 5 shows the spectrum of 19.2% methanol/80.8% CO2 (30mL of methanol in the syringe) doped into the plasma. The OH band intensity increased significantly compared to the 6.5% methanol case. The intensities of the all CN and C2 bands decreased even more. The hydrogen emission line at 486.1 nm was intense. This line was not observed with 6.5% methanol, but was observed with 19.2% methanol. Comparison of Figures 4 and 5 illustrates the significance of the effects of mobile-phase composition on spectral background. For example, with the lower methanol composition, observation of atomic emission in the 306.0-nm region might be possible. However, the increase in molecular band emission as the methanol concentration is increased to 19.2% might make these observations impossible. Exactly the opposite may be said of the 436.5-nm spectral region. Effect of Pressure on Spectral Background with the Binary Mobile Phase. Figures 6-8 show the spectra of 10%
400
600
Wavelength (nm) Flgure 6. Spectrum of 10 % methanol/90 % C02 Introduced into the helium discharge. The frtt restrictor was used with a pressure of 250
atm.
I
200
I
I
400
600
Wavelength (nm) Figure 7. Spectrum of 10% methanol/90% C02 Introduced into the helium dlscherge. The frit restrlctor was used with a pressure of 200
atm.
methanoV90% COZfrom a Scott Specialty Gas SFC grade mixture. The frit restrictor with pump pressures of 150,200, and 250 atm were used. In many respecta, the 250-atm spectrum (Figure 6) is similar that of Figure 5. Minor differences include the observation that the helium lines at 587.6,501.57, and 447.1 nm are intense in Figure 6. The CN bands at 359.0 and 388.3 nm are much less intense. The bands from C2 and NH as well as the carbon line are similar in intensity. In general, the remainder of the molecular bands is reduced in intensity. As the SFC pressure is reduced, the decreased mass flow of the mobile phase into the plasma
ANALYTICAL CHEMISTRY, VOL. 65, NO. 22, NOVEMBER 15, 1993
9293
#
0
2
N
3
2,4'dlohl-one
'5 w L
3
2
6
4
7
0
Time (minutes)
200
I
I
400
600
Flgure
@.Chromatographic separation with the Integral restrictor.
Column temperature was 120 O C .
Wavelength (nm) Fbure 8. Spectrum of 10 % methanol/90 % C02 introduced Into the helium discharge. The frlt restrlctor was used with a pressure of 150
atm.
causes the intensities of the OH, CN, and CZmolecular bands to decrease. This behavior may be seen by examination of Figures 7 and 8. Background Spectra Examination Summary. While the results of these experiments produced much information, the important details with regard to the spectral background can to be reduced to a few major points. In the absence of the methanol modifier,the spectral background from the COZdoped helium plasma in the UV/visible spectral region consists largely of atomic emission from carbon and molecular bands from Cz,CN, and NH. Methanol has a dramatic effect on spectral background, giving rise to an intense OH band. The OH band intensity increased dramatically as the methanol concentration was increased. As the methanol concentration is increased in the experiments with 0 and 6.5 7% methanol, the CZband emission becomes less intense for the lower energy transitions and more intense for more higher energy transitions. It is possible that the CZis coupling with the higher energy OH moiety. As the concentrationis increased to 19.8% methanol, the intensities of all CZbands decrease. It is likely that plasma cooling dominates in the latter case. In general, the CN molecular bands became less intense when methanol was introduced. These 200-600-nm background emission studies for the SFC mobile phases have shown that the MIP can be employed as an element-selective detector which is compatible with methanoVCOz binary mobile phases. However, care must be taken to avoid spectral interferences. Chlorine Emission Line Examination. Chlorine emission was characterized with a Raoult's law based sampling device.16 The sample solution was 7 pL of a 4.5 % (v/v)solution of CHzClz in squalane, and the helium flow through the sampler was 50 mL/min. The most intense chlorine emission lines in the UV/visible spectral region are at 479.45,481.01, and 486.13 nm. These lines reside in the region close to the CZbands. Fortunately, the most intense line at 479.45 nm did not overlapthese molecular emission bands. The presence of COZintroduced with the SFC pump at a pressure of 250 atm quenched chlorine emission by roughly 50%. Effect of Helium Flow Rate on Chlorine Emission Intensity. The effect of helium support gas flow rate on emission intensity for chlorine was examined in the flow range from 12 to 17 L/min. Chromatograms were obtained with 107% methanol in carbon dioxide mobile phase using the 7-10 mL/min integral restrictor. Injections of 140 ng of 1,3dichlorobenzene were monitored. At 12 L/min, the chlorine emission intensity was greatest. The peak areas decreased (16)Webster, G.K.;Carnahan, W.W.A d . Chem. 1989,61,790-793.
2,4'dichlorobenzophenone
1,ddichlorobenzene
hexachlorobenzene
l L 0
1
2
3
Time (minutes) Chromatographlc separation with the Integral restrictor. Column temperature was 130 O C . linearly to 56% of this value as the flow rate was increased to 15.5 L/min. The lower helium flow rate gave longer analyte residence time in the plasma region producing more efficient atomization and excitation. However, due toincreased plasma noise at these flow rates, the worst signal-to-noiseratio was exhibited at lower flow rates. This observation is probably caused by a larger perturbation of the plasma by COz at lower He flows. The ratio of COz to He increases as the He flow decreases. All subsequent chromatograms were obtained at flows of 16 L/min. SFC Mobile-Phase Pressure Effects on Chlorine Emission Intensity. The effects of COZ mobile-phase pressure on peak area were examined for 2,4'-dichlorobenzophenone,p-chlorobiphenyl, and hesachlorobenzene in the range of 150-300 atm in 50-atm increments. In general, peak areas increased as the pressure was increased from 150to 250 atm. An increase of pressure from 250 to 300 atm resulted in a decrease in peak areas. Trends detailed in the literature indicate that increased SFC mobile-phase loading in the Flgure 10.
3294
ANALYTICAL CHEMISTRY, VOL. 65, NO. 22, NOVEMBER 15, 1993
plasma decreases analyte emission.8*gJlJ2 This behavior was seen only at higher pressures. It is possible that increases in peak areas as the pressures were increased from 150 to 250 atm were due to improved analyte transport as the presswe and, consequently, the velocity of the analyte leaving the restrictor increased. For experiments which follow, the pressure is maintained at 250 atm. Chromatograms. A series of chromatogramswas obtained under various conditions. Examples of these with the 10% methanol mobile phase and the 7-10 mL/min integral restrictor are shown on Figures 9 and 10. Figure 9 shows the separation of three nonvolatile compounds, hexachlorobenzene, p-chlorobiphenyl, and 2,4'dichlorobenzophenone, with benzene as the solvent. The column temperature was 120 "C. Figure 10 shows the separation of 1,3-dichlorobenzene, hexachlorobenzene, and 2,4'-dichlorobenzophenone with tetrahydrofuran as the solvent with a column temperature of 130 OC. Compared to Figure 9, the retention time was reduced and the peaks were more Gaussian in shape. It should be noted that in earlier SFC/MIP experiments with the nonpolar pure COZmobile phase, Zhang et al.11 and Webster and Carnahan12 found detection of higher molecular weight, later eluting compounds such as hexachlorobenzene to be difficult. These works were with lower" and moderate power12 plasma systemswith column pressures of 80-250 atm. A major advantage in the reduced retention times was seen in this work. The faster retention times result primarily from the use of the binary mobile phase and the increased SFC pressure. The differences in the retention times in Figures 9 and 10 are significant. With the benzene solvent, a longer retention time resulted. Although not shown, these longer retention times occurred with the benzene solvent at both 110 and 130
"C. It is possible that the increased retention times with benzene are caused by phenyl stationary-phase-benzene/ analyte interactions which increase k' for the phenyl-type analytes examined here. CalibrationPlots and Detection Limits. Mass detection limits were determined as the amount of analyte which yielded a peak height 3 times the standard deviation of the chromatographic baseline. Relativedetection limitswere obtained by dividing mass detection limits by the peak widths. The detection limits were 12 ng or 1.9 ng/s for p-chlorobiphenyl, 15 ng or 1.4 ng/s for 2,4'-dichlorobenzophenone,and 19 ng or 1.6 ng/s for hexachlorobenzene. Calibration studies for p-chlorobiphenyl, 2,4'-dichlorobenzophenone, and hexachlorobenzene were performed in the range of tens of nanograms to 400 ng. All compounds yielded linear response over -2 orders of magnitude with lower ranges roughly a factor of 2 greater than the detection limit.
CONCLUSIONS The MIP has proven to be an effective detector for SFC compatible with methanol-modified COz mobile phases. Significant analysis advantages accrue with the shorter retention times produced by the binary mobile phase. Nonvolatile polar chlorinated compounds can be detected effectively with reasonable linear dynamic ranges and detection limits. It is important to consider spectralbackground effects as the introduction of additional components alters these spectra. Both the compounds of the binary phase and their relative compositions must be considered. RECEIVEDfor review June 14, 1993. Accepted August 20, 1993." ~
~
~
Abstract published in Advance ACS Abstracts, October 1, 1993.
