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Anal. Chem. 1980, 52, 2278-2282
Combined Capillary Gas Chromatography/Atmospheric Pressure Negative Chemical Ionization/Mass Spectrometry for the Determination of 2,3,7,8-Tetrachlorodibenzo-p-dioxin in Tissue R. K. Mitchum," G. F. Moler, and W. A. Korfmacher Department of Health and Human Services, Food and Drug Administration, National Center for Toxicological Research, Jefferson, Arkansas 72079
By use of a Capillary column gas chromatograph interfaced directly to an atmospheric pressure Ionization mass spectrometer, the concentratlon of 2,3,7,8-tetrachlorodibenzo-pdioxin (TCDD) was determined in environmentally exposed fish and wildlife. Isolation of TCDD from tissue samples was accomplished via multistep high-pressure liquld chromatography Incorporating stable label Isotope dilutlon. Polychlorinated biphenyls were found not to interfere with TCDD analysls at the low parts-per-trillion levels.
T h e choice of analytical techniques employed for the (TCDD) resanalysis of 2,3,7,8-tetrachlorodibenzo-p-dioxin idues has received a n ever-increasing amount of attention because of the possible toxic effects stemming from environmental exposure to TCDD ( I , 2). T h e mass spectrometric determination of TCDD a t the parts-per-trillion (10-l' g/g) level depends upon the concentration of contaminants (3, 4 ) such as 1,1'-(2,2,2-trichloroethylidene)bis(4-~hlorobenzene) (DDT), a minor component of toxaphene, and polychlorinated biphenyls (PCBs) which interfere with the analysis. PCBs have been found to interfere with both the low- a n d high-resolution mass spectral measurements of TCDD (3). Therefore, it is important that the sample cleanup portion of the analytical procedure be as selective as possible for TCDD. T h e sample cleanup procedures currently being used in other laboratories have been reviewed by Lamparski and Nestrick ( 5 ) ,who recently have developed a multistep high-performance liquid chromatography (HPLC) method to separate and isolate synthetic isomeric TCDDs (6). T o date the mass spectrometric techniques for identifying a n d quantitating these toxic residues are based upon ions derived from the electron impact mass spectrum of TCDD under either low- or high-resolution conditions (7-9). Hass et al. ( 1 0 , I I ) have investigated the behavior of polychlorinated dibenzo-p-dioxins by use of negative chemical ionization mass spectrometry (NCI/MS) employing a methane/oxygen plasma at 1-2 torr (100-250 Pa) working pressure. T h e sensitivity of their method precluded the analysis of environmental levels of TCDD, but the results were consistent with the earlier work by H u n t et al. (12) for the reaction of the negative ion, 2,3,7,8-tetrachlorodibenzo-p-dioxin, with 0 2 . Atmospheric pressure ionization mass spectrometry (API/MS) has been found t o be a sensitive analytical technique having a high degree of selectivity due to the thermal nature of the ion molecule collisions and the long ion source residence times (13,14). The application of the API technique for the analysis of TCDD utilizing a n oxygen-rich plasma should take advantage of both the sensitivity of the API and the selectivity of the negative ion molecule reaction explored by H u n t e t al. (12) and developed by Hass et al. (11). In this work we describe a new method for the quantitation of TCDD in nonfat tissue samples based upon the addition
of a stable label isotope diluent followed by a new multicolumn HPLC cleanup procedure and capillary gas chromatography/negative ion atmospheric pressure ionization/mass spectrometry (GC/NIAPI/MS).
EXPERIMENTAL SECTION Apparatus. G C f N I A P I I M S . The atmospheric pressure ionization mass spectrometer was fabricated on contract by Extranuclear Laboratories, Inc. (Pittsburgh, PA), and has been described previously (15, 16). The system consists of an atmospheric pressure ionizer utilizing a 1-mCi 63Ni foil as the primary electron source. The source is flange mounted on a differentially pumped vacuum chamber incorporating two 800 Lfs turbomolecular pumps (Airco Temscal Model 814). The front chamber houses the low-pressure ion optics which incorporate a filament. The rear section houses the quadrupole mass filter (Extranuclear) and continuous dynode electron multiplier assembly (Gallieo Model 4516, Sturbridge, MA). As shown in Figure 1, the ion source has been modified to interface to a HewlettPackard Model 5710 gas chromatograph. The capillary injector consists of a (Chrompack)1-mm i.d. glass capillary insert adapted to fit a modified Finnigan Model 9500 GC aluminum block injector assembly. The glass capillary column (20 m Supelco SP-2100 SCOT) was connected to the glass injector with Teflon shrink tubing (Analabs, 0.8 mm). The column outlet was connected directly to the atmospheric pressure ionizer via a 0.12 mm i.d. Pt/Ir tubing (Chrompack 6521) transfer line. The nitrogen carrier flow rate was 8 cm3/min, measured with a (Brooks 5841-1AlAC) flow meter. The GC conditions used were as follows: injection port, 250 "C; column temperature, hold for 2 min and then program from 65 to 220 OC at 32 OCfmin. The effluent of the capillary column was made up with a nominal 10% oxygen (Linde ultra high purity)/nitrogen mixture prior to ionization. The purified nitrogen was obtained from liquid nitrogen boil off after passing through 13X molecular sieves which had previously been baked at 400 "C. The inlet gas manifold was made of stainless steel with welded bellows sealed valves throughout (Nupro SS-4BRG). Those experiments which required an oxygen-free system utilized an "Oxytrap" (Alltech 4003) in series with the gas purification system. As a confirmation of the oxygen-free status of the system, the spectrum of 1-chloro-4-nitrobenzenewas taken to determine the extent of phenoxide ion formation (14). The ratio of negative molecular ion to phenoxide anion was typically 3:1, indicating a nearly oxygen-free carrier gas (14). The output of the electron multiplier was capacitive coupled to a pulse amplifier-shaper-discriminator(Extranuclear, Inc.). This signal was fed into a dedicated Incos minicomputer system (Finnigan Corp., Model 2400) which had been modified to be operated in the pulse counting mode. The selected ion monitoring (SIM) data were recorded by using a f0.5 amu window for each peak with an integration time of 0.21 sfwindow. The two masses that were monitored for TCDD and TCDD-13C12,respectively, were m f z 176 and 182. The resolution was sufficient to obtain base-line separation. Electron Capture Gas Chromatography Conditions. The column used for the electron-capture gas chromatography (EC/GC) was 5 m X 4.0 mm i.d. with 3% OV-101 on 80/90 mesh Anakrom Q packing. The carrier gas was P5 (5% methane in argon), set at a flow rate of 30 mL/min. The column was operated
This article not subject to U.S. Copyright. Published
1980 by the
American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 52, NO. 14, DECEMBER 1980
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Figure 1. GC/NIAPI/MS inlet schematic: (1) injector; (2)glass capillary adaptor; (3)Teflon shrink tubing; (4) 20-m capillary column; (5)glass to metal tubing seal; 6) Pt/Ir capillary; (7)quartz injector tube; (8) source volume; (9) Ni foil; (10) 50-wrn aperature; (1 1) vacuum housing; (12)GC oven wall; (13)makeup gas inlet; (14)overflow gas outlet.
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isothermally at 240 "C, and the detector temperature was 350 "C. Reagents. TCDD. The isotope diluent, uniformly ring-labeled 2,3,7,8-tetrachlorodibenzo-p-dioxin-13C,2 (TCDD-13C),was prepared on contract (Midwest Research Institute, Contract No. (synthesized by 222-76-2037[c]) from 2,4,5-tri~hlorophenol-'~C~ Los Alamos Laboratories under Interagency Agreement No. FDA-IAG-74-36[0]). The isotopic purity was determined from the 70-eV electron impact spectra to be 97.8 atom % I3C with an isotopic distribution of 78.4% 13C12,17.4% 13Cll,and 4.2% 13C10. The unlabeled TCDD was obtained from ECO-Control, Inc. (Cambridge, MA), and was used as received. Preparation of Chromatographic Columns. The polyurethane foam-harcoal packing material was prepared by using the method of Huckins et al. (17). The foam-charcoal column was prepared by dry-packing this material into an 8 cm X 4.2 mm i.d. stainless steel HPLC column. The silica gel column was prepared by slurry packing (18) 5 pm silica gel (Spherisorb S5W, Laboratory Data Control [LDC], Riviera Beach, FL, catalog no. 615027) into a 30 cm X 3 mm i.d. glass-lined stainless steel HPLC column (LDC catalog no. 600029). The reversed-phase (RP) column was prepared by slurry packing (18)5 pm ODS material (Spherisorb S5 ODS, LDC catalog no. 625036) into a 30 cm X 3 mm i.d. glass-lined column. All solvents were either Burdick and Jackson "distilled-in-glass" or Mallinckrodt Nanograde, except for the 95% ethanol (Pharmco DSP grade). Other chemicals were ACS grade, Procedure. The sample cleanup consists of five steps. The sample was homogenated and then subjected to digestion, extraction, foam-charcoal chromatography, silica gel HPLC, and reversed-phase HPLC. The quantitation was via GC/NIAPI/MS. Homogenation and Digestion. A 50-g sample of tissue and 200 mL of deionized water (DW) were homogenized by using a Polytron homogenizer (Brinkman Instruments, Westburg, NY). The homogenate was transferred to a 750-mL Erlenmeyer flask. One hundred milliliters of 95% ethanol and 50 ng of TCDD-13C [l part-per-billion (ppb)] were added to the homogenate, which was stirred for 1 h. Seventy-five milliliters of NaC1-saturated water and 50 mL of freshly prepared (hot) KOH-saturated water were then added to the flask. The solution was stirred for 15 min. Another 50 mL of hot KOH-saturated water was added, and the solution was stirred for another 15 min. Extraction. One hundred milliliters of hexane was added to the flask, and the mixture was stirred briskly for 15 min. The hexane phase was then transferred to a 500-mL separatory funnel, and the homogenate was reextracted three additional times with 100-mL portions of hexane. The hexane extract was washed with 100 mL of a 10% solution of NaCl in DW and then with four successive 100-mL portions of DW. The hexane extract was then passed through an anhydrous Na2S04column (2.54 cm x 5 cm
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plugged with unsilanized glass wool). Foam-Charcoal Chromatography. The hexane extract was pumped through a foam-charcoal column by using a metering pump (Labpump Model RP-SY-lCSC/R479, Fluid Metering, Inc., Oyster Bay, NY). The column was then washed with 100 mL of hexane followed by 50 mL of benzene/ethyl acetate (50/50 mixture). The direction of the column flow was then reversed and the TCDD-containing fraction eluted with 150 mL of toluene, the eluate being collected in a 200-mL beaker. The beaker was placed on a hot plate in a hood, and the eluate evaporated to near dryness under a stream of air. The concentrate was transferred to a 2-mL reaction vial equipped with a Teflon-lined cap. The concentrate was slowly taken to dryness with high purity N2 and then redissolved in 25 pL of CHC13for injection onto the silica gel HPLC column. Silica Gel HPLC. The 25 WLCHC13solution was injected onto the silica gel column, and the fraction of the eluate, corresponding to the retention time window of TCDD, was collected in a 2-mL reaction vial. TCDD-W was used to determine the proper collection time for both RP and silica gel HPLC. TCDD-13Cwas employed for this purpose to avoid possible cross-contamination of the tissue samples. TCDD eluted with the solvent front by using a hexane flow rate of 1.0 mL/min. The silica gel column was cleaned with CH3CN between sample runs. Reverse-Phase HPLC. The fraction from the silica gel column was taken to dryness with purified N2 and redissolved in 7 pL of CHC13. This CHC13solution was then injected onto the reversed-phase HPLC (RP-HPLC) column, and a fraction of the eluent corresponding to the retention time of TCDD was collected in a 2-mL reaction vial. The eluent was acetonitrile/water (7525) flowing at 1.0 mL/min. The acetonitrile was pesticide grade and the water had been extracted three times with pesticide grade hexane. TCDD eluted at about 9.5 min under these conditions. The R P column was cleaned with CHC1, between sample runs. Sample Preparation. The TCDD fraction from the RP-HPLC column was evaporated to dryness with purified nitrogen and the residue dissolved in hexane for EC/GC and GC/NIAPI/MS analysis. The sample was typically concentrated to 10 pL and a l-lrL portion injected onto the GC/NIAPI/MS. Sample Quantitation. Quantitation was performed by obtaining the ratio of the peak areas for unlabeled TCDD and TCDD-W for a sample and comparing this to a standard curve. The standard curve was obtained by GC/NIAPI/MS analysis of mixtures of unlabeled TCDD and TCDD-13Cstandards under the same instrumental conditions as the sample analyses.
RESULTS AND DISCUSSION T h e reaction of 2,3,7,8-tetrachlorodibenzo-p-dioxin with oxygen in a n electron-rich plasma has been investigated by Hunt et al. (12) and later by Hass e t al. (10) using conventional low-pressure (1-2 torr) negative ion sources. T h e conclusion drawn from their work is consistent with eq 1 and proposes
the attack of the molecular radical anion of TCDD upon the neutral Oz molecule. T h e support for this reaction is the observance of the TCDD molecular negative ion in the absence of O2and the observance of the 4,5-dichloro-l,2-benzoquinone radical anion in the presence of Oz, I n an oxygen-rich plasma t h e major negative ions a t atmospheric pressure are formed from both electron capture and clustering processes according to eq 2. T h e abundance of
e-
+ O2 + Nz
-
Oz- + N2
these negative ions is a function of both the ion source temperature and the concentration of O2in t h e nitrogen buffer gas. Under the conditions used in this work, the NIAPI/MS
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ANALYTICAL CHEMISTRY, VOL. 52, NO. 14, DECEMBER 1980
176
178 m/z
180
Figure 2. Resolved mass spectrum of the 4,5dichlor~l,24enzoquinone anion formed via reaction of TCDD with 02-.
reagent ion spectrum consisted of 02-, OzN2-,04-,and 02N4in an intensity ratio of 100:6:36:3,respectively. T h e NIAPI mass spectrum of TCDD in an oxygen-rich plasma is given in Figure 2. Under these conditions, the negative ion mass spectrum of unlabeled TCDD is shown to be dominated by the negative ion a t m / z 176, the 4,5-dichloro-1,2-benzoquinoneradical anion. The molecular negative ion of TCDD was not detected. In addition, under the same conditions, but without the oxygen reagent gas, the molecular negative ion of TCDD was still not observed contrary to the findings of Siege1 et al. (15) and Reynolds et al. (16). At atmospheric pressure in the absence of electric fields the electron energy distribution is Maxwellian and is a function of the temperature of the system. T h e failure of TCDD to electron capture under these conditions indicates that TCDD does not have a maximum in its electron capture cross section near thermal energies. The production of molecular negative TCDD ions in experiments a t low pressure (12) is consistent with the capture of nonthermal electrons which are in abundance in these ion sources. The transition state for the ion molecule reaction in eq 1 is symmetric and does not prevent the reaction of the oxygen negative ion with neutral TCDD. Therefore, at atmospheric pressure, under thermal conditions, eq 3 is thought to be operable.
By the GC/NIAPI/MS technique, it was found that the standard curve for the peak area ratios vs. the TCDD-13C/ TCDD (weight/weight) injected ratio was linear from a concentration ratio range of 1:l to 9O:l. Because the T C D D - W concentrations were not corrected for isotopic purity nor were they calculated on a molarity basis, the slope of this standard curve should be approximately equal to the fraction of the TCDD-13C standard that was fully ring-labeled (assuming no instrumental quantitation artifacts). I n fact, if the slope of the standard curve is calculated on a molarity basis, the slope is 0.76, and the fraction of the TCDD-13C standard that was fully ring labeled was 0.78, vide supra. T h e final phase of the cleanup procedure is RP-HPLC. Figure 3 shows sequentially run RP-HPLC chromatograms of a TCDD-l3C standard and a TCDD-l3C spiked tissue extract, cleaned up through the silica gel step. Samples collected from the RP-HPLC step could be examined by EC/GC for cleanliness and for total TCDD (labeled plus unlabeled) levels. Figure 4 shows the chromatograms obtained from EC/GC analysis of a TCDD-13Cstandard and a cleaned u p tissue sample. T h e total recovery was determined by EC/GC which provided a measurement of the absolute amount of TCDD and TCDD-13Cin the isolated sample. This information, combined
Retention Time (min.)
Figure RP-HPLC 3ce of: (a) TCDD-I3C s ndard; (b) fish flesh extract initially spiked with 1 ppb TCDD13C. The conditions are listed in the Procedure section.
1 1 1
EC/GC
j
I I
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0
4
8
1
2
Retention Time (mi..)
Figure 4. EC/GC trace of: (a) 100 pg of TCDO-13Cstandard; (b) turtle egg and liver extract initially spiked with 1 ppb TCDD13C. The conditions are listed in the Procedure section.
with the capillary GC/NIAF'I/MS measurement of the ratio of the TCDD to TCDD-13Cin the sample, and the knowledge of the TCDD-13C spike level, was used to determine the percent recovery in these samples. The recovery of TCDD-13C varied, ranging from 5 to 50%. While part of the variation in recovery may be due to differences in the sample matrices, a like range was found for a set of similar fish tissue samples. Due to limited sample size, multiple analyses were not performed, so an assessment of either the sample matrix effects or the efficiency of the subsequent cleanup procedure could not be determined. While low in absolute value, a recovery range of this magnitude
ANALYTICAL CHEMISTRY, VOL. 52, NO. 14, DECEMBER 1980
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Figure 5. GC/NIAPI/MS response obtained for a standard mixture of 60 pg of TCDD and 600 pg of TCDD-13C: (a) response at m / z 176; (b) response at m l z 182.
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il 0
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400
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800
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Figure 6. GC/NIAPI/MS response for a fish (predator)extract spiked with 1 ppb TCDD-13C. This result corresponds to a level of 15 ppt TCDD in the sample. is not uncommon in the determination of TCDD in environmental samples. However, preliminary results have shown that substituting benzene for hexane as the extracting solvent resulted in higher TCDD recoveries. In addition, the advantage of the isotope dilution method used in this procedure is that it corrects for losses of the analyte that may occur during the sample isolation steps. Therefore, as long as the standard curve is well-defined for the ratio of the ion chromatogram peak areas vs. the ratio of the amounts of the analyte and its isotopic diluent, analyte quantification is greatly facilitated. Thus, absolute recovery values need not be high, as long as the total amount recovered is sufficient for detection. The detection limit, however, will increase as the absolute recovery decreases since the limit of detection of the mass spectrometer is fixed. Caution must be exercised since substrate binding to the matrix components would result in data variability which is not compensated for by addition of a stable label isotope diluent. Figure 5 shows the selected ion chromatograms obtained from injection onto the GC/NIAPI/MS of a standard hexane solution containing 60 pg of TCDD and 600 pg of TCDD-13C. Figure 6 shows the selected ion chromatograms for a fish extract spiked a t 1 ppb with TCDD-13Cwhich was found to contain 15 ppt TCDD. Figure 7 shows similar data obtained from another fish tissue extract and corresponds to a level of 230 ppt TCDD. Figures 6 and 7 demonstrate the ability of the procedure's multicolumn cleanup method to separate TCDD from the sample matrix and to provide quantitative data. Similar results were obtained from a variety of other tissue matrices including frog muscle and liver, turtle eggs and liver, snake muscle, and squirrel muscle and liver (Table I).
2 3 4 5 6 7 8 9
10 11 12 13 14
tissue typea,b turtle egg and liver fish (edible flesh) snake liver and muscle catfish muskrat liver racoon liver buffalo fish frog liver and muscled fish (predator) fish (predator) fish (bottom feeder) catfish buffalo fish
catfish
PPtC 145
480 60 40
ND (40) ND (10) ND ( 1 3 ) >10 000 230 15 77 50 ND ( 7 ) ND ( 7 )
a All samples were obtained from two rivers in Arkansas: The Arkansas River and the Bayou Meto (a tributary of the Arkansas River). All samples were spiked at 1 part-per-billion ( p p b ) with TCDD-I3C. ND = none detected; the number in parentheses is the measured detection limit for that sample; ppt = parts-per-trillion. Sample was obtained from a holding pond behind Vertac. Inc.. Jacksonville. AR.
The detection limit varied with the recovery for these samples but was on the order of 10-30 ppt for most of the (50 g) samples. The detection limit was taken as the concentration of TCDD required to give a signal 2.0 times the noise level. The detection limit obtained in this procedure results from the combination of two factors. The first is that a relatively large sample (50 g) was analyzed. The second is that the absolute detection limit of the GC/NIAPI/MS was low for TCDD (approximately 10 pg), using the selected ion monitoring technique to take advantage of the inherent sensitivity of the atmospheric pressure ionization source. The high selectivity of this procedure is obtained by the combination of three parameters. The first is the cleanup procedure which separates out many possible interfering compounds. The second is the capillary GC separation portion of the GC/NIAPI/MS analytical step. The third is the use of the unique negative chemical reaction (eq 3) and selected ion monitoring of only the ions of interest. One class of possible interferences with TCDD determination by mass spectrometric methods is the presence of PCBs in the sample. T o check for this possible interference in the quantitation of TCDD, we analyzed Arochlor mixtures by the
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formation of the phenoxide anions under atmospheric pressure ionization conditions from chlorinated aromatic hydrocarbons has been observed in our laboratory as well as by Dzidic (20). Therefore, by analogy, on the basis of the above findings, monitoring of the phenoxide anion at m / z 301 could result in a differentiation between 2,3,7,8 and 2:2 substituted tetra-CDD isomers having a peri chlorine. However, until the other 12 2:2 substituted isomers can be tested by the proposed method, the ability to distinguish 2,3,7,8-tetrachlorodibenzo-p-dioxin from these 12 other isomers is uncertain. In this experiment the combination of HPLC cleanup and capillary GC precludes the detection of dioxins other than tetra-CDD isomers on the basis of retention time. The 20-m capillary column used in this experiment did not have the efficiency to separate all 13 of the 2:2 substituted tetra-CDD isomers. Although this experiment was not designed to quantitate individual tetra-CDD isomers, we are presently developing methodology based upon a 60-m capillary column which should fulfill these requirements.
ACKNOWLEDGMENT T h e authors wish to thank J. Spohn and D. Firestone for providing the initial fish extracts for TCDD analysis and W. Holtman for supplying samples of contaminated fish and wildlife. We also wish to acknowledge K. Rowland for his assistance in some of the sample cleanup steps.
LITERATURE CITED (1) Gasiewicz, T. A.; Neal, R. A. Toxicol. Appl. Pharmacol. 1970, 57, 329-339. (2) Barsotti, D. A.; Abrahamson, L. J.; Allen, J. R. Bull. Environ. Contam. Toxicol. 1079, 21, 463-469. (3) Baughman, R.; Meselson, M. A&. Chem. Ser. 1973, No. 720,92-104. (4) O'Keefe, P. W.; Meselson, M. S.;Baughman, R. W. J. Assoc. Off. Anel. Chem. 1078, 6 1 , 621-626. (5) Lamparski, L. L.; Nestrlck, T. J.; Stehl, R . H. Anal. Chem. 1979, 57, 1453-1458. (6) Nestrick, T. J.; Lamparski, L. L.; Stehl, R . H. Anal. Chem. 1970, 57, 2273-2281. (7) Hummel, R. A.; Shadoff, L. A. Anal. Chem. 1980, 52, 191-192. (8) Kimble, 8. J.; Gross, M. L. Science 1980, 207, 59-81. Wilklnson, M. K.; Dupuy, A. E., Jr.: (9) Harless, R. L.; Oswald, E. 0.; McDaniel, D. D.; Tai, H. Anal. Ch8m. 1980, 52, 1239-1245. (10) Hass, J. R.; Frlesen, M. D.; Harvan, D. J.; Parker, C. E. Anal. Chem. 1978, 50, 1474-1479. (1 1) H a s , J. R.; Friisen. M. D.; Hoffman, M. K. Org. Mass Spectrom. 1070, 74, 9-16. (12) Hunt, D. F.; Harvey, T. M.;Russel, J. W. J. Chem. SOC.,Chem. Commun. 1975, 151-152. (13) Horning, E. C.; Horning, M. G.; Carroll, D. I.; Dzidlc, I.; Stillweli, R. N. Anal. Chem. 1973, 45, 936-943. (14) Horning, E. C.; Carroll, D. 1.: DzMic, I.; Lin, S.-N.; Stillwell, R. N.; Thenot, J.-P. J. Chromatogr. 1977, 742, 481-495. (15) Siegel, M. W.; McKeown, M. C. J . Chromatcgr. 1078, 722, 397-413. (16) Reynolds, W. D.; Mitchum, R. K.; Newton, J.; Byskoff. R. I.; Pomeynack, C.; Brand, H.; Siegel, M. W. Chem. Instrum. (N.Y . ) 1977, 8, 63-98. (17) Huckins, J. N.; Stalling, D. L.; Smith, W. A. J. Assoc. Off. Anal. Chem. 1878, 61, 32-38. (18) Nomura, A.; Morka Y.; Kogure, Y. Bunseki Kagaku 1078, 2 7 , 504-509. (19) Buser, H. R.; Rappe, C. Chemosphere 1978, 7, 199-211. (20) Dzidic, I.; Carroll, D. I.; Stillwell, R. N.; Horning, E. C. Anal. Chem. 1075, 47, 1308-1312.
RECEIVED for review May 19,1980. Accepted September 23, 1980. Presented in part at the 8th International Conference on Mass Spectrometry, Oslo, Norway, 1979.
