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Anal. Chem. 1983, 55,2232-2236
Automated Molecular Absorption Spectrometry for Determination of Fluorine in Biological Samples Pothapragada Venkateswarlu,* Larry D. Winter, Robert A. Prokop, and Donald F. Hagen] Commercial Chemicals Diuision/3M, Building 236-GB53, 3M Center, St. Paul, Minnesota 55144
Methods descrlbed so far for determining fluorine by aluminum monofluoride molecular absorption spectrometry using the graphlte furnace call for Injection of the aluminum matrix solution, drying, and ashlng, followed by a separate Injection of the sample. These two-Injection procedures wlth intervening drying and ashing steps are not adaptable to autosampling. The present procedure Is simpler and requires only one injection of the sample sultably dlluted with the matrix solution. This procedure Is directly amenable to automation and saves considerable amounts of time and labor. Evidence Is presented to show that not all types of organlc fluorine compounds can be analyzed for fluorine by molecular absorption spectrometry foilowlng direct Injection of the sample Into the furnace. This shortcoming has now been overcome through the use of the sodium biphenyl reagent to convert organlc fluorine Into inorganic fluoride before the Injection of the sample Into the furnace.
The aluminum monofluoride molecular absorption procedures for fluorine determination described by Tsunoda et al. (I), Chiba et al. (8,and Fujimori et al. (3) involve two separate injections into the graphite furnace, one of the aluminum matrix solution and another of the sample, with specific temperature programs after each injection. Such two-injection procedures with separate temperature-time programs cannot be carried out by the autosampling devices now furnished with the atomic absorption spectrophotometers. On the basis of careful study of the various factors governing the generation of the aluminum monofluoride absorption signal, a new procedure has now been developed that is simpler, requires only one injection, and can be automated to save time and labor. This procedure is described in this paper. Chiba et al. (2) reported that the serum fluorine values obtained by the molecular absorption method were significantly higher than the ionic fluoride (inorganic) values they obtained by direct measurement with the fluoride ion selective electrode and were also similar to the total fluorine values (inorganic plus organic F) obtained after ashing the samples. On the basis of these observations, they confirmed the earlier reports on the two fractions of fluorine (inorganic and organic/covalent) in blood serum (4,5)and concluded that their method is suitable for direct determination of total fluorine (inorganic and covalent) in serum without any sample treatment. Fujimori et al. (3) have also reported a similar aluminum monofluoride molecular absorption method for determining total fluorine in serum. The difference between the total fluorine and the inorganic fluoride would reflect organic fluorine in the serum samples. If the inorganic fluoride value is insignificant relative to total fluorine, the value SO obtained by the molecular absorption method would essentially reflect organic fluorine in the serum samples. Because of our interest at 3M in rapid screening of blood serum samples from plant workers for organic fluorine (6, 7), we investigated the applicability of the molecular absorption method proposed by Chiba et al. (2)for direct determination Central Research Laboratories/3M.
of organic fluorine in blood serum samples, without any preliminary treatment. The results so obtained, however, were significantly lower than those obtained with the sodium biphenyl method or the oxygen bomb-reverse extraction method reported by Venkateswarlu (8, 9). This problem of low recoveries of total fluorine with the blood serum samples was now corrected through the use of the sodium biphenyl reagent whereby the organic fluorine is converted into inorganic fluoride prior to the injection of the sample into the furnace.
EXPERIMENTAL SECTION Reagents. Reagent grade chemicals and double distilled deionized water were used throughout. Sodium fluoride stock solution, 1000 pg/mL F, contained 2.2105 g of sodium fluoride in 1L of water. Organic fluorine stock solution, 1000 pg/mL F, contained 1.5128 g of ammonium perfluorooctanoate (3M Co.) in 1 L of water. These stock solutions were suitably diluted to obtain fluorine standard solutions of desired strength covering the range of fluorine in the samples. The aluminum matrix solution I contained 0.1 M aluminum nitrate (37.513 g of Al(N0&9HzO/L of water), 0.05 M nickel nitrate (14.541 g of Ni(N03)2.6H20/Lof water), and 0.05 M strontium nitrate (10.582 g of Sr(NO3I2/Lof water). Suitable volumes of this stock matrix solution were added to the sample to provide an aluminum ion concentration of 0.01 M in the final sample solution injected into the furnace. The stock matrix solution was diluted 10-fold to obtain aluminum matrix solution 11. Sodium biphenyl reagent, flash point (closed cup) 30 "C, was obtained from Southwestern Analytical Chemicals, Inc. Perchloric acid (Mallinkrodt) used was 70%. Diphenylsilanediol (DPSD) solution was prepared by dissolving 2 g of DPSD (Pierce) in 1 L of toluene. Apparatus. The equipment used included a Perkin-Elmer absorption spectrophotometer (Model 4000) with the background corrector, platinum lamp (303-6051),furnace programer (HGA400), standard graphite furnace (290-1633), and autosampler (AS-40) and multitube vortexer (Scientific Manufacturing Industries, Model 2600). The 5-mL polypropylene stoppered tube, Falcon tube 2063 used in our earlier studies (8), is now being manufactured by a different process and the stopper leaks. A combination of Falcon tube 2053 with Falcon cap 2032 was found to be leakproof and was used for processing samples in these studies, as well as in all our current fluorine analyses of samples by the sodium biphenyl-fluoride electrode procedure (8). Procedures. (A) Inorganic Fluoride. The sample was diluted if necessary to contain fluoride in the range 0.1-0.5 pg of F/mL. To 900 p L of the diluted sample was added 100 pL of aluminum matrix solution I. Inorganic fluoride standards (0.0, 0.1,0.2,0.3,0.4, and 0.5 fig of F/mL) were also similarly diluted with the aluminum matrix solution I. Samples and standard solutions so diluted with the aluminum matrix solution were injected into the furnace and absorbances at 227.5 nm measured. Details of injection volumes and conditions for drying, ashing, atomization, and measurement of absorbance are indicated under program D in Table I. The fluoride contents of the samples are determined from the calibration curve obtained with the standard solutions. ( B )Organic Fluorine. To 20 p L of the sample (blood serum) in a 5-mL polypropylene test tube (Falcon tube 2053 fitted with Falcon cap 2032) was added 500 pL of ether, followed by 5 p L of perchloric acid. The extraction of the fluorochemical from the sample into the organic liquid phase was achieved by vigorous vortexing of the contents for 15 min or better by placing the
0003-2700/83/0355-2232$01.50/00 1983 American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 55, NO. 14, DECEMBER 1983
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Table I. Furnace Programmings Employed by Various Workers for Analysis of Fluorine by Aluminum Monofluoride Molecular Absorption Spectrometry program D program C program B a program A' ref 3 present method ref 2 ref 1 steps involved matrix plus sample 25 p L matrix soln matrix soln matrix soln injection I 20 p L 20 p L 10 pL 150 "C 25 A (400 "C) 230 "C 20 A (330 "C) drying I 30 s/ramp 20 s 30 s/ramp 10 s 20 s hold 700 "C 60 A (850 "C) ashing I 10 s/ramp 15s 10 s hold stop injection I1 drying I1 ashing I1 atomization and measurement
cool furnace
cool furnace
cool furnace 1 5 s
sample 5 ML 20 A (330 "C) 10s 40 A (640 "C) 30 s 280 A (3300 "C) 7s
serum 5 p L 25 A (400 "C 20 s
serum 5 p L 230 "C 30 s/ramp 830 "C 60 s/ramp 3000 "C 7s
60 A (850 "C) 15s 280 A (3300 "C) 7s
2200 "C 0 s/ramp 10 s-hold stop gas flow
a Temperature was indicated by the current applied to the furnace: 70 A = 1000 "C and 200 A = 2400 "C. Temperatures in parentheses are approximate extrapolations.
sample in an ultrasonic bath for 15 min. Two milliliters of sodium biphenyl reagent was added and the tube promptly stoppered. During a 10-min reaction time the contents were vortexed or placed on a shaker. The sodium biphenyl reagent was destroyed by adding 0.5 mL of water. The contents were vortexed vigorously for 10 min to extract the fluoride into the aqueous layer and centrifuged at 2500 rpm for 2 min. The supernatant was aspirated and the aqueous layer washed twice with ether (0.5 mL each time). To the aqueous extract was added 1mL of diphenylsilanediol (DPSD) solution followed by 1 mL of ice-cold perchloric acid. Extraction of fluoride as diphenyldifluorosilane (DPDFS) into the organic phase was carried out by vortexing the tube for 10 min. After centrifugation of the tube at 2500 rpm for 5 min, 750 pL of the DPDFS layer was transferred to another tube (Falcon 2053 tube, 2032 cap) containing 100 p L of concentrated ammonium hydroxide. Reverse extraction of the fluoride ions into the aqueous layer was accomplished by vortexing the contents for 10 min on the multiple vortexer. After centrifugation of the tube, the supernatant organic phase was removed by aspiration. The aqueous phase was washed twice with heptane (0.5 mL) and diluted with 400 p L of water. The reverse extracts, so diluted, were transferred to the autosampler which was programmed to add 50 p L of aluminum matrix solution I1 to 50 p L of the reverse extract injected into the furnace (before commencement of drying I step, program D Table I). Organic fluorine standard solutions (0, 1,2,3,4,5 pg of F/mL) were also processed as above simultaneously with the samples. The absorbance was measured, and the fluorine content of the samples was calculated as described in the case of inorganic fluoride samples.
RESULTS AND DISCUSSION (A) Simplification of the Molecular Absorption Procedure and Its Adaptation to Autosampling. All the programs, A, B, and C (Table I), call for a preliminary injection of the aluminum matrix solution, drying, and ashing, followed by the injection of the sample and a repetition of the drying and ashing cycles prior to atomization and measurement of the absorbance. Tsunoda et al. (1)did not explain clearly the basis for recommending in their procedure two separate injections, one for the matrix solution and another for the sample, with an intervening drying step, beyond stating: "An excess aluminum solution is applied first to the furnace before the fluorine containing solution is pipetted, so that the A1F molecule is effectively formed inside the furnace". Such a
0.~1
400
600
800
1000
1200
1400
1600
1800
Temperature " C
Flgure 1. Effect of ashing temperature on the observed aluminum monofluoride absorbance. Five microliters of 1 pg/mL of fluoride in aluminum matrix solution
I1 was injected each time.
two-injection procedure is not amenable to autosampling. To save time and labor in analysis, the feasibility of adapting this procedure to autosampling was now explored. The effect of ashing temperatures on the aluminum monofluoride absorption signals from the matrix plus fluoride solution was investigated. The findings are shown in Figure 1. The decline in the absorbance signal beyond the 800 "C ashing temperature suggested to us that the (aluminum) fluoride molecules are swept out of the furnace during ashing at temperatures exceeding 800 "C before the absorbance is measured. Therefore, if the ashing of the sample could be completed well within the temperature range in which the fluoride molecules are not so lost, it should be possible to accurately measure the fluorine in the sample following a single injection of the sample plus matrix solution. This indeed was found to be the case. The procedure described in this paper prescribes a single injection of the sample suitably diluted with the aluminum matrix solution, followed by drying (150 "C), ashing (700 "C), atomization (2200 "C), and measurement of absorbance. This procedure is amenable to autosampling and saves a considerable amount of labor and time. In the case of both the procedures, A and B, the two-injection procedures described earlier by others require over 8
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ANALYTICAL CHEMISTRY, VOL. 55, NO. 14, DECEMBER 1983
Table 11. Accuracy and Precision of Determination of Fluoride by the Aluminum Monofluoride Molecular Absorption Method (Procedure A, 25 pL Injections) MmL of F expected 0.05
&mL of F found mean f std dev
% re1
n
error
std dev
-4
0.06
0.048 0.060
U.08
0.081 i. 0.0044
* 0.0014
7 7 6
0.10
0.100+ 0.0027 0.21Of 0.0040 0.300 i- 0.0040 0.380 It 0.0040 0.510f 0.0160
8 8 8 8 8
0.20
0.30 0.40
0.50
i
0.0016
% re1
3.3 2.3 5.4 2.7 2.1 1.4
0
+1.3 0 t5 0
-5 1-2
1.0
3.1 ~~
~~
h of continuous attention of the analyst for triplicate runs of 35 samples plus standards (one manual injection approximately every 2 min). The present one-injection automated procedure requires no more than half an hour of the operator’s time to load the autosampler with the same number of samples and standards. (13) Inorganic Fluoride. Inorganic fluoride in solutions of samples can be determined by procedure A, described in the Experimental Section. Inasmuch as the molecular absorption method for fluorine is extremely rapid and sensitive, paradoxically it has its limitations too. Unlike the fluoride electrode procedure, the molecular absorption procedure cannot be used for direct determination of inorganic fluoride in the presence of covalent fluorine unless the organic fluorine compound is volatile enough to be swept out of the furnace before it could react with the matrix to form aluminum monofluoride. Otherwise, the covalent fluorine is partly or completely converted into inorganic fluoride which is indistinguishable from the native inorganic fluoride in the sample. The accuracy and precision of the method for analysis of inorganic fluoride (procedure A), over the range 0.05-0.5 pg of F/mL and using 25-pL injections, were investigated (Table 11). The results were accurate to within 5% of the expected values and precise to within 5% relative standard deviation. The “characteristic concentration” for the method, which is a measure of sensitivity expressed as nanograms of analyte corresponding to 1%absorption or 0.0044 absorbance, seemed to vary slightly from furnace to furnace from the same lot. The best value that we obtained was 0.04 ng of F compared to 0.021 ng of F and 0.028 ng of F reported by Tsunoda et al. (1)and Fujimori et al. (3). The sensitivity did not change on switching from the one-injection procedure to the two-injection procedure. (C) Organic Fluorine in Biological Samples. Need for Cleaving the Covalent Fluorine Bonds. Samples of blood serum of plant workers handling fluorochemicals were analyzed directly by the aluminum monofluoride molecular absorption method (2) and also by the sodium biphenyl/fluoride electrode method (8). In the case of several samples, the results obtained by the molecular absorption method were lower than those obtained with the sodium biphenyl method (Table 111). Variable degrees of loss of organic fluorine during the ashing of the samples in the graphite furnace was suspected. However, Tsunoda et al. (1) have earlier reported quantitative recoveries of fluorine from sodium monofluoroacetate, trifluoroacetic acid, and o-fluorobenzoic acid by direct molecular absorption spectrometry, and they indicated that their method may be suitable for total fluorine determination without any sample treatment. We investigated recoveries of organic fluorine, in the case of a few selected fluorochemicals, by the same molecular absorption method they described. The recoveries varied from 100 to 0.0% (Tables IV and V). Apparently, during some phase of the heating of the furnace some organic fluorine molecules or fragments thereof
Table 111. Comparison of Results of Total Fluorine Analysis of Blood Serum Samples of Fluorochemical Plant Workers
sample
total F, pg/mL sodium biphenyl, directAlF fluoride electrode molecular absorption 2 5 I 9 11 12 13 17
Table IV. Recovery of Fluorine in Selected Organic Fluorine Compounds by Direct Molecular Absorption Spectrometry
compound Freon E-2 C,HF,,O, Freon E-5 C,,HF,,O, FOMB YO-4 (C,F,O), 1H, 1H, 2H, 2Hperfluorooctanol lH,lH,2H,Wperfluorodecanol perfluorodecanoic acid 1-fluoronaphthalene p - fluoroanisole perfluorotributylamine perfluorodecalin dibromoperfluorobutane
ngof ngof Fa F % injected found recovery 12.5 12.5 12.5 12.5
3.1 8.0 8.1 3.8
29.6 64.0 64.8 30.4
12.5
2.7
21.6
12.5 12.5 12.5 12.5 12.5 12.5
3.8
30.4
0.0
0.0
4.6
36.8
0.0 0.0 1.1
0.0 0.0 8.8
a Water extracts of the compounds were analyzed for covalent fluorine by the sodium biphenyl/fluoride electrode method and suitably diluted to contain 12.5 ng of F per 25-rL injection,
are swept out of the furnace prior to i;teraction of the fluorine with the aluminum matrix to form aluminum monofluoride. The molecular absorption method is, however, an exceedingly convenient method for determining inorganic fluoride. T o take advantage of this technique and to obtain reliable results for covalently bound fluorine in organic compounds or biological samples, it was decided to convert the organic fluorine in the samples to inorganic fluoride by use of the sodium biphenyl reagent as was done in the case of determining covalent fluorine with the fluoride ion electrode (8). The fluoride ions obtained following cleavage of the covalent bonds by the sodium biphenyl reagent were extracted into a small volume of water. This extract is highly alkaline and detrimental to the formation of the aluminum monofluoride molecules in the furnace and so the extract was neutralized. However, the high salt content and the organic matter (from biphenyl), both of which did not interfere with the fluoride ion electrode measurement, did interfere with the molecular absorption measurement. There was spattering and smoking in the furnace resulting in erroneous absorbance signals. The extract was then submitted to the reverse extraction technique described by Venkateswarlu (10) for concentration and separation of fluoride free from interferences. Reverse extract obtained with sodium hydroxide contained organic matter in amounts lesser than before, but which yet caused problems. Finally, the reverse extraction with ammonium hydroxide yielded a preparation with much less organic matter and was satisfactory for analysis by molecular absorption spectrometry. This was the basis of the procedure B described in this paper. The reverse extraction technique was explored by use of trimethylchlorosilane and diphenylsilanediol. Although the
ANALYTICAL CHEMISTRY, VOL. 55, NO. 14, DECEMBER 1983
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Table V. Comparison of Results of Total Fluorine Analyses of Blood Serum Samples Spiked with Various Amounts of Selected Fluorochemicals total F, bg/mL Na biphenylserum direct mol abs mol abs expected sample fluorochemical added a b
ammonium perfluorooctanoate
C
(N-ethylperfluorooctanesu1fonamido)ethy1 alcohol
d e
potassium perfluorooctanesulfonate
f g h i j
k 1
0.4 1.0 2.0 3.0 0.3 0.6 1.0 1.9 4.4 7.3 12.5 13.3
4.0, 4.6 9.4, 9.4 14.8, 14.8 16.9, 16.9 5.0, 3.5 9.8, 10.6 13.7, 14.3 25.7, 24.9 4.6, 4.9 9.1, 9.6 12.1, 13.4 17.3, 17.2
4.9 9.8 14.6 19.3 5.0 12.5 16.6 25.0 4.4 8.6 12.6 16.6
Table VI. Comparison of Results of Total Fluorine Analyses of Blood Serum Samples from Plant Workers
serum sample no.
direct mol abs
a b
4
C
4 5 4 8 7 9 10
d e f g
h i
total fluorine, pg/mL Na biphenyl/ oxygen bomb/ reverse reverse extr-mol abs extr-mol abs
1
recoveries of covalent fluorine were the same in both cases (procedure B), the molecular absorbance of “no-fluorine”blank solutions was larger with trimethylchlorosilane (0.5 absorbance) than that with diphenylsilanediol (0.05 absorbance). Hence diphenylsilanediol was preferred to trimethylchlorosilane for reverse extraction of fluoride in procedure B. Several 2O-pL aliquots of a solution of ammonium perfluorooctanoate (3 pg/mL F) were analyzed by procedure B, using 50-pL injections (0.0045 pg of F). The results (F in pg/mL) were 3.05 mean f 0.165 standard deviation ( n = lo), with a relative error of 1.7% and a relative standard deviation of 5.4%. Table V contains results of fluorine analysis by the present method of serum samples to which variable amounts of selected organic fluorine compounds were added. These results are also compared with those obtained by direct molecular absorption method proposed by Chiba et al. (2). Except in the case of potassium perfluorooctanesulfonate,the recoveries by the direct molecular absorption method were inadequate. Essentially quantitative recoveries were obtained by the same procedure following intervention with the sodium biphenyl reagent. These results stress the need for conversion of organic fluorine to inorganic fluoride prior to analysis by the molecular absorption method. Chiba et al. (2)found that the fluorine values of three serum samples obtained with the molecular absorption method (0.13,0.12, and 0.21 pg of F/mL) were significantly higher than the inorganic or ionic fluoride values obtained with the fluoride electrode (0.076,0.025, and 0.02 pg of F/mL, respectively) and concluded that the molecular absorption method measures total fluorine (inorganic and organic F) in serum samples without any preliminary treatment. In support of their conclusions, they furnished results of total fluorine analyses of the very same serum samples, following open ashing, by the molecular absorption procedure (0.16, 0.21, and 0.27 wg of F/mL) and with the
5 5 8 9 11 13 15 22 23
6 7 8 9
18 23
Na biphenyl/ fluoride electrode 5 5 6 7 8
9 12 18 20
fluoride electrode (0.24, 0.18, and 0.29 pg of F/mL, respectively). They considered that these values, although slightly higher, were essentially similar to those obtained earlier (0.13, 0.12, and 0.21 pg of F/mL) by direct molecular absorption spectrometry without any pretreatment of samples. They attributed the slightly higher values obtained with the ashing methods compared to those obtained by the direct molecular absorption measurement to possible contamination of the samples with extraneous fluoride during open ashing. The chances for contamination of samples in the graphite furnace would be far less. While they considered the possibility of loss of volatile organic fluorine compounds in the molecular absorption technique, they overlooked the possibility of loss of organic fluorine in the open ashing procedure as well (9, 12). Therefore, while the results obtained by both the procedures could be similar, the recoveries of covalent fluorine need not necessarily be 100% in both the cases. It would have been preferable if they had supported their conclusions with results of total fluorine analysis of serum samples obtained by procedures not susceptible to errors due to contamination or loss of fluorine as may occur in open ashing. These errors are not encountered in the oxygen bomb (9) and sodium biphenyl (8)decomposition methods, that were employed in the present investigations. Table VI contains results of such total fluorine analysis of actual serum samples from plant workers handling fluorochemicals. The results obtained by direct molecular absorption spectrometry are significantly lower than those obtained by other methods involving the conversion of organic fluorine to inorganic fluoride, viz., sodium biphenyl/fluoride electrode, sodium biphenyl/reverse extraction/molecular absorption, and oxygen bomb/reverse extraction/molecular absorption methods. There is a satisfactory agreement among the total fluorine results obtained by the latter three methods. These results demonstrate partial loss of covalent fluorine in direct molecular absorption
2236
Anal. Chem. 1983, 55,2236-2238
spectrometry as in the conventional open ashing procedure and once again bear out the need for complete conversion of organic or covalent fluorine in serum samples to inorganic fluoride prior to injection of the sample into the furnace for total fluorine determination by molecular absorption spectrometry. Results obtained by the above procedure measure total fluorine in serum. Inorganic fluoride is most conveniently measured directly with the fluoride ion electrode or in the case of microsamples by the hanging drop electrode technique (11). The difference between the total fluorine and the inorganic fluoride would be the value for covalent fluorine in the sample. If the inorganic fluoride concentration is insignificant, total fluorine and covalent fluorine concentrations are essentially the same. This method (procedure B) has not, so far, been used by us to determine organic fluorine in normal human or animal blood sera, in which the organic fluorine levels would be relatively lower than those in the samples from plant workers exposed to organic fluorochemicals. However, following extraction of organic fluorine from larger samples (2-5 mL, instead of 20 pL employed in the present procedure), it is possible to attain organic fluorine levels that fall within the analytical range of the molecular absorption method. Such an approach has been described earlier by one of us (8).
ACKNOWLEDGMENT We wish to thank Wanda Bahmet for her valuable help and discussions in the early phase of these investigations. Thanks
are also due to Gary W. Kirsch, Mark A. LaCroix, Barbara A. Coil-Nelson, and Sheila D. Kromer for excellent technical help. Registry No. F, 16984-48-8;F2,7782-41-4; sodium biphenyl, 5137-46-2; Freon E-2, 3330-14-1; Freon E-5, 37486-69-4; FOMB YO-4, 25038-02-2; 1H,1H,2H,2H-perfluorooctanol,647-42-7; lH,lH,W,W-perfluorodecanol, 678-39-7;perfluorodecanoic acid, 335-76-2;1-fluoronaphthalene, 321-38-0;p-fluoroanisole, 459-60-9; perfluorotributylamine, 311-89-7; perfluorodecalin, 306-94-5; dibromoperfluorobutane, 73533-18-3; ammonium perfluorooctanoate, 3825-26-1; (N-ethylperfluorooctanesu1fonamido)ethyl alcohol, 1691-99-2;potassium perfluorooctanesulfonate,2795-39-3.
LITERATURE CITED (1) Tsunoda, K.; Fujlwara, K.; Fuwa, K. Anal. Chem. 1977, 49, 2035-2039. ( 2 ) Chiba, K.; Tsunoda, K.; Haraguchi, H.; Fuwa, K. Anal. Chem. 1980, 52, 1582-1585. (3) Fujimorl, S.; Itai, K.; Sakurai, S.; Tsunoda, H. Fluoride, in press. (4) Taves, D. R. Nature (London) 1968, 277, 1050-1051. (5) Venkateswariu, P.; Singer, L.; Armstrong, W. D. Anal. Biochem. 1971, 42, 350-359. (6) Ubel, F. A.; Sorenson, S. D.;Roach, D.E. Am. Ind. Hyg. Assoc. J . 1980, 41, 584-589. (7) Beiisie, J.; Hagen, D. F. Anal. Blochem. 1978, 8 7 , 545-555. (8) Venkateswarlu, P. Anal. Chem. 1982, 5 4 , 1132-1137. (9) Venkateswariu, P. Anal. Biochem. 1975, 68, 512-521. (10) Venkateswarlu, P. Anal. Chem. 1974, 46, 878-882. (1 1) Venkateswarlu, P. Clln. Chlm. Acta 1975, 59, 277-282. (12) Kakabadse, G. J.; Manohln. J. M.; Bather, J. M.; Weller, E. C.; Woodbridge, P. Nature (London) 1971, 229, 626-627.
RECEIVED for review April 18, 1983.
Accepted September 1,
1983.
Determination of the Isotopic Composition of Oxygen by Electron Spin Resonance Spectrometry Nis Bjerre* Institute of Physics, Aarhus University, DK-8000 Aarhus C, Denmark Elfinn Larsen Chemistry Department, Riser National Laboratory, DK-4000 Roskilde, Denmark
The isotopic composition of gaseous oxygen at -0.5 torr is determined by measuring the relative intensities of the distinct ESR signals from "02,160180, and I8O2and converting the Intensities to amounts of the isotopic species by means of intensity factors derived from the transition moments and Boltzmann factors. The precision and linearity of the ESR method are investigated with samples of oxygen prepared by thermal decomposltlon of KBrO, of known Isotopic composition and, furthermore, the method Is compared with mass spectrometry. For micromole samples of natural isotopic composltlon (0.2% I6O), the "0 content can be measured wlth a relatlve preclslon of 2%, whereas the preclslon obtained with enriched samples is better than 1%.
In connection with studies of the photolysis of crystalline halates ( I ) , we encountered the problem of determining the isotopic composition of molecular oxygen emanating slowly
from leO-enriched crystals exposed to ultraviolet light. We found that this analysis could conveniently be carried out by irradiating the crystals in an evacuated and sealed quartz tube and monitoring the ESR signals of the oxygen gas produced. The ESR spectrum of gaseous oxygen in the electronic ground state, 32;, reflects the coupling between the electronic spin and the rotational angular momentum (2,3). Accordingly, the spectra of the different isotopic species are distinct as apparent from Figure 1. In this paper we describe how the isotopic composition of gaseous oxygen can be determined by measuring the relative intensities of selected lines in the ESR spectrum. The method is compared with a mass spectrometric analysis.
EXPERIMENTAL SECTION Materials. The oxygen gas was prepared by thermal decomposition of solid KBr03. The decomposition was carried out on platinum or gold since it was found that considerable isotopic exchange occurs if KBr03 is decomposed in contact with glass or quartz. The KBr03 of "natural" isotopic composition was
0 1983 American Chemical Society 0003-2700/83/0355-2236$01.50/0
