Anal. Cham. 1989, 61, 408-412
408
Partial support for this research provided by the National Science Foundation under Grant CHE-85-19087 to the University of California, Riverside, is gratefully ac-
(18) Here, P. J. J. Magn. Reson. 1983, 55, 283-300. (19) Ha, S. T. K.; Lee, R. K. L; Wilkins, C. L. J. Magn. Reson. 1987, 73,
6, 1988.
467-476.
Received for review September 12,1988. Accepted December
knowledged.
Fluorometric Measurement of Aqueous Ammonium Ion in Flow Injection System
a
Zhang Genfa1 and Purnendu K. Dasgupta* Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, Texas 79409-1061
column derivatization method in regard to how long before chromatography the derivatives must be prepared. In looking for more stable derivatives amenable to electrochemical detection, Jacobs (12) recently found that, sulfite, in lieu of ME, offers advantages. The reaction can also be used for the sensitive detection of sulfur dioxide (13). reaction for Although Taylor et al. (14) used the OPAthe determination of ammonia shortly after the discovery by Roth, subsequent exploitation of this reaction has primarily been focused on the determination of amino acids. The measurement of ammonia and ammonium ion by this reaction is enjoying a resurgence in the present decade (15-19) culminating in a recent study by Goyal et al. (20) which attempts the optimization of all pertinent parameters of the OPA-ME reaction, specifically for determining NH3/NH4+. We have discovered that the spectral characteristics of the OPAsulfite-NH3 reaction product are considerably more attractive than the corresponding product with ME, both for absorptiometric or fluorometric detection. Here we present the details of this method, as adapted to a flow-injection system.
An automated flow-injection method for NH3/NH4+ Involving the ternary reaction of the analyte with o-phthaldialdehyde (OPA) and sulfite Is described. The use of malodorous thiol compounds Is avoided and the reaction provides much greater sensitivity by either fluorescence (detection limit 300 fmol or 20 nM NH4+) or absorption detection compared to the reaction Involving 2-mercaptoethanol. The reaction shows considerable selectivity for ammonia over amino adds by a factor of 16 to >500 for 11 common amino acids studied. The
throughput rate Is 25 samples/h.
Ammonia is the principal atmospheric base responsible for the neutralization of atmospheric acidity (1). Ammonium salts, most notably sulfates, are frequently the principal components of the submicrometer fraction of the atmospheric aerosol burden (2) and are widely believed to be the primary responsible agents for the degradation of atmospheric visibility (3). Indeed, ammonium bisulfate, not organic compounds, was shown to be the dominant species in fine particle aerosol in the Great Smoky Mountains (4). Sensitive determination methods for NH3/NH4+ are needed to improve the time resolution of atmospheric measurements. Classically, ammonia has been determined by the indophenol blue reaction (limit of detection (LOD) 10 jug/L) or Nessler’s reaction (LOD 20 Mg/L) (5). In 1971, Roth discovered the ternary reaction of o-phthaldialdehyde (OPA), a “reducing agent” (borohydride or mercaptoethanol (ME)), and ammonia or primary amino acids to produce intensely fluorescent products and described its analytical usefulness (6). Detailed studies have shown the generality of the reaction and the products with thiols (which behave as nucleophiles, not reducing agents) have been unequivocally characterized to be l-(alkylthio)isoindoles (7, 8). Either through precolumn derivatization or postcolumn conversion, this reaction has become the major basis of chromatographic analysis of amino acids, and has appropriately become a textbook example (see, e.g., ref 9). Like many other ternary reactions, the reaction can be used to determine any of the three components involved. Thiols have been the analytes of interest for Nakamura and Tamura (10) and Mopper and Delmas (11). The latter authors also showed that sulfite and sulfide react in the same fashion as thiols. The (alkylthio)isoindole derivatives are not especially stable and present limitations in the pre-
EXPERIMENTAL SECTION Reagents. Standard grade o-phthaldialdehyde (P-1378, Sigma Chemical Co., St. Louis, MO) was used in this work without further purification. Experimentation with in-house twice recrystallized OPA or a “specially purified grade” available from the supplier did not show any significant benefits over the standard product. The OPA reagent is prepared by dissolving 268 mg of OPA, in 50 mL of methanol, followed by the addition of 150 mL of water. The solution, 10 mM in OPA, can be stored refrigerated for 1 week. We do not recommend inclusion of buffering agents in the OPA preparation, in our experience this decreases useful life of the reagent and leads to variable blanks. Phosphate buffer (0.1 M) is made by dissolving 26.81 g of analytical reagent grade Na2HP04 in 900 mL of water, adjusting pH to 11.0 with 2 M NaOH and diluting to 1 L. Sodium sulfite solution (3.0 mM, 0.378 g/L) is prepared daily in the phosphate buffer. Carbonate and borate buffers used (see results) were in the form of the sodium salt. Ammonium standards were made from a 0.1000 M NH4C1 solution. Extreme care is necessary to make low-level standards, freshly deionized water must be used and exposure to ambient air avoided. Such standards must be used immediately after preparation. Analytical System. The apparatus arrangement is schematically shown in Figure 1. A peristaltic pump P (Minipuls 2, four-channel head, Gilson Mediad Electronics, Middleton, WI) is used to pump water (W) as carrier at 50 ^L/min through a short column C (55 X 3 mm poly(tetrafluoroethylene) (PTFE) tube packed with monosize H+-form cation exchange resin (Bio-Rex MSZ 50, Bio-Rad Laboratories, Richmond, CA), glass wool retaining plugs, replaced weekly) to remove traces of NH3/NH4+ and then through an electromechanically actuated six-port rotary
1 Permanent address: Shanghai Hygiene and Anti-Epidemic Center, 280 Chang Su Rd., Shanghai, People’s Republic of China.
0003-2700/89/0361-0408Í01.50/0
©
1989 American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 61, NO. 5, MARCH p
---OPA-ME,pH6.8
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Excitation (nm)
Figure 1. System schematic: P, pump; B, buffered sulfite; 0, ophthaldlaldehyde; W, water; C, H+-form cation exchanger microcolumn; 1, 2, knotted mixing coils; V, rotary loop injection valve; S, sample; R, heated reactor, thermostated at 85 °C; T, porous tubing; F, fluorescence detector.
HVXL 6-6, Hamilton Co., Reno, NV). The injection loop constituted a 124 X 0.3 mm PTFE tube; the measured injection volume (which included significant contributions from valve internal volume) was 14 ^L. The sample S is aspirated by the pump through the valve. A chronTrol CD-4S timer (Lindberg Enterprises, San Diego, CA) was utilized to automate the load/inject functions of valve V. The carrier stream merges with the OPA reagent stream 0 (50 /uL/min) at a low-volume tee, is followed by a knotted mixing coil Ml (0.3 x 250 mm), and merges again with the buffered sulfite reagent stream B (50 gL/min). The next mixing coil M2 (0.3 X 1000 mm) is followed by a heated stainless steel reaction coil R (0.5 x 500 mm) kept in a water bath thermostated at 85 ± 1 °C. (For the determination of high NH4+ levels, the heated reactor is omitted altogether, vide infra.) The elevated temperatures invariably result in bubble formation in the flow stream; these are removed by a short segment (1-2 cm) of a porous hydrophobic membrane tube T (Gore-Tex 001, W. L. Gore and Associates, Elkton, MD) prior to entry into the fluorescence detector F, aided by a small amount of flow restriction at the detector exit (21). The fluorescence detector (Fluoromonitor III, Laboratory Data Control, Riviera Beach, FL) was equipped with a 351-nm phosphor-coated 1-in. mercury pen lamp (BHK, Inc., Monrovia, CA), a broad band-pass (approximately 280-380 nm) glass excitation filter (Oriel Corp., Stratford, CT), and a long-pass emission filter (50% cutoff at 425 nm) and utilized a 30 mL volume flow cell. All connections were made with 0.3 mm i.d. PTFE tubing. Liquid reservoirs were protected from the intrusion of ammonia by acidic traps; we found acid-washed silica best suited for this purpose. Absorption and ratio-corrected fluorescence spectra were obtained with a spectrophotometer (Model 559) and a spectrofluorometer (Model MPF-448, both from Perkin-Elmer), respectively. valve V (type
RESULTS AND DISCUSSION of the Reaction Product and Characteristics Spectral Detection Sensitivity. The absorption and emission spectra of the OPA-sulfite-NH3 reaction product is shown as the solid trace in Figure 2. The excitation spectra (not shown) are essentially the same as the respective absorption profiles. Similar spectra for the OPA-ME-NH3 reaction product are shown as dashed and dotted traces, respectively, following two recent published procedures (19,20) for reagent compositions and relative volumetric ratios. The final solution for all three cases contained the same concentration of ammonium, 100 µ each for the absorption spectra and 10 µ each for the emission spectra. The spectra shown represent a reaction time that yields the maximum amount of product under each condition; for the present sulfite method and one of the ME procedures (20), this required controlled heating (60 °C, 2 min). For the other ME procedure (19), maximum fluorescence was observed after 10 min at room temperature. All reactions were conducted in 0.1 M phosphate buffer. The emission spectra (obtained in each case at the excitation maximum) clearly show the superior fluorescence properties of the sulfite reaction product. Note that both the absorption and emission spectra for the reaction product with sulfite are shown on a 3-fold reduced ordinate. The substitution of a more acidic sulfonic acid group for a thiol functionality is
Figure 2. Absorption (a) and emission (b) spectra of products of OPA-NHg-sulfite (pH 11.0), OPA-NHg-Me (pH 6.8), and OPA-NHg-ME (pH 9.5). See text for details.
expected to lead to a greater dipole moment for 1sulfonatoisoindoles relative to 1-(alkylthio)isoindoles. The red shift of the absorption maximum (as measured for the product formed under alkaline conditions) and the greater absorptivity of the sulfonic acid derivative may be thus explained. Large increases in emM and red shift of the Xmal in going from the methylthiol to the methylsulfonic acid derivative are well-known for p-aminotriphenylmethane dyes, e.g., pararosaniline (22). The stronger fluorescence of the 1sulfonatoisoindole appears to largely result from increased absorptivity. The optimum excitation/ emission conditions (Xeimax 365 nm, Xemmal 425 nm) was not utilized for the actual analytical system due to lack of appropriate components at the time the majority of work was conducted (the long pass filter with 50% cut off at 425 nm cuts off nearly half of the emitted light). A study of detector optical parameters was subsequently made. Of various excitation source/filter combinations, a simple “black light” (miniature tube, General Electric F4T5) in combination with a 360-nm excitation filter and a 420-nm long-pass emission filter, provided the best sensitivity, more than twice that obtained under the conditions stated above.
Reaction Kinetics: Effect of pH, Buffer Composition and Temperature. The OPA-ME-NH3 reaction under alkaline conditions (19) is fast, it is typically complete in under 2 min at room temperature. At neutral pH (20) the reaction is much slower but provides greater discrimination against amino acids (14). Sulfite is a weaker nucleophile than a thiol and as such the reaction is expected to be slower for sulfite. Under the operating conditions (residence time in the reactor ~40 s at 85 °C), the effect of varying the pH (the pH of the 0.1 M phosphate buffer is cited, the reaction pH is only marginally different) is shown in Figure 3a. The optimum pH, 10.5-11.0, is similar to that reported by Jacobs (13) albeit his study involved high concentrations of amino acids as test samples. The time necessary for the completion of the reaction with NH4+ under typical trace analysis conditions is significantly higher than that reported by Jacobs. The optimum pH is also virtually the same as that for the reaction involving ME (10, 16,17). Interestingly, the reaction kinetics are dependent not only on pH but also on the choice of the buffer. Figure 3b shows that the rate of fluorescence development (t = 22 °C) is quite
different for phosphate, carbonate and borate buffers, all adjusted to pH 11.0 and having a total concentration of 0.1 M. Specific acid catalysis from buffering ions is not uncommon, given the same total buffer concentration, often the rate depends on the (relevant) pK of the buffering ion (23). This trend is also true in the present case, phosphate (pK3 12.4) buffers perform better than carbonate (pK2 10.3) which
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8
40
I
20
0
0
Sulfite Concentration OPA Concentration
Buffer
pH
Reaction
Time
(mini
Figure 4, Effect of reagent concentrations on fluorescence Intensity: (a) dependence on OPA concentration; (b) dependence on sulfite
concentration. Test NH4+ concentration was
Figure 3. (a) Effect of reaction pH on fluorescence intensity, phosphate buffer; (b) effect of the choice of buffering agent on reaction kinetics (22 °C), 0.1 M, pH 11.0 In all cases (A, phosphate; B, carbonate; C, borate); (c) comparison of system output with the three buffers In b; (d) effect of reactor temperature (residence time 40 s), 0.1 M phosphate, pH 11.0. Total residence time of sample in the system for all cases was
~110
s.
performs significantly better than borate (pK 9.2). This behavior is also reflected in actual performance in the flow-injection system at room temperature, as shown in Figure 3c. In the mechanism suggested for the reaction of OPA, ME, and primary amines (8), at least one of the steps involved should be subject to acid catalysis. We therefore conducted some exploratory experiments in which the reaction was allowed to proceed first in acidic conditions and then made alkaline, or the reaction was conducted in an alkaline-acidalkaline sequential regime. While none of these efforts resulted in accelerated formation of the desired product, some unexpected observations were made; we believe that they are worthy of mention. When the OPA reagent and sulfite were added to the ammonium sample, followed by an acidic buffer (pH 0-2), a purple product appeared immediately. The compound showed two distinct absorption bands at 430 and 540 nm of approximately equal absorptivity at pH 0; the 430-nm band dominated by a 3:1 ratio at pH 2. This product was nonfluorescent or very weakly fluorescent. It converted slowly to the product with Xmu 365 nm (the fluorescent product) in acidic solution and much more rapidly in alkaline solution. Inasmuch as sulfite was necessary for its formation, it could not simply be phthalimidine, the spectral characteristics also rule out a phthalimidine. Interestingly, the product was not produced under similar conditions with ME, nor was it produced when acid was first added to the sample, followed by OPA and sulfite. Experiments with other primary amino compounds showed lack of a similar reaction with n-pentylamine, leucine, alanine, arginine, asparagine, glutamic acid, cysteine, and tyrosine while ethanolamine (Xm„ 480 nm), aniline (450 nm), lysine (495 nm), glycine (495 nm), and tryptophan (575 nm) reacted and the respective products displayed only one absorption peak as indicated. While not highly sensitive, this reaction, not previously described in the literature, may conceivably be utilized for selective determinations. While manipulation of pH was not fruitful, increasing the reaction temperature (phosphate buffer, pH 11) was effective
(mM)
(mM)
1
µ.
in accelerating the desired fluorogenic reaction. The effect of the reactor temperature on the analyte response is shown in Figure 3d. Goyal et al. (20) also utilized an elevated reaction temperature for the OPA-ME-NH3 system at neutral pH, but unlike that study we did not observe a decreased response at elevated temperatures. This may be due to the greater stability of the sulfonato derivative (12). Effect of Reagent Concentrations. With 1 µ NH4+ as the test sample, the effect of OPA concentration in the reagent was determined (other conditions as in the Experimental Section). The results, shown in Figure 4a, prompted us to choose an OPA concentration of 10 mM; this choice is not markedly different from the optimum values recommended by others (11,20) for the thiol reaction system. The base-line fluorescence value is directly related to the OPA concentration and therefore it is not desirable to use an OPA concentration greater than 10 mM. Similarly, the effect of sulfite concentration was studied with other parameters held constant as specified in the Experimental Section. On the basis of the results shown in Figure 4b, a 3.0 mM concentration of sulfite was chosen. The sulfite concentration does not appear to have any significant effect on the system blank. The relative independence of the response on the precise sulfite concentration near 3 mM allows for some oxidative decay of the sulfite solution to occur before any decrease in response is observed. Nevertheless, we find it advisable to prepare the sulfite solution daily. The use of thiosulfate, which is considerably more stable than sulfite, was found to produce a much lower fluorescence signal, under otherwise comparable conditions. To obtain the best detection limits, we find it essential to keep the OPA and sulfite reagents separate. Attempts to combine the two reagents result in high and variable blank levels and typically show persistent base-line drift (increasing blank) during the course of the experiment. It is possible that the combined reagent blank increases from the slow intrusion of ammonia, or that a fluorogen reported to be formed slowly at room temperature between OPA and sulfite (24) is responsible for this behavior. Effect of Sample Salinity. Goyal et al. (20) made the unexpected observation that the response to a NH3/NH4+ sample in their analytical system was dependent on the ionic strength. We therefore tested the effect of up to 0.1 M NaCl and 0.1 M Na2S04, individually, upon the response due to 5 µ NH4+. The blank response due to these salts at 0.1 M concentration indicated the presence of 0.09 and 0.16 µ NH4+ in these salts, i.e., 0.27 and 0.20 ppm NH4+ was present, respectively, in the original reagents. Except for this blank response, no effects of salinity could be discerned; the analyte signal due to 5 µ NH4+ did not change upon the addition of the salts up to concentrations indicated above. System Performance. On the basis of the signal to noise ratio (S/N) observed for a 50 nM sample, the detection limit, according to the criteria specified by the American Chemical Society Committee on Environmental Improvement (25), S/N
ANALYTICAL CHEMISTRY, VOL. 61, NO. 5, MARCH
relative
relative sample
ammonia alanine arginine asparagine cysteine glutamic acid
response 100.00 6.02 0.30 5.07 0.28 1.22
± 0.89 · 0.20 ± 0.02 ± 0.01 ± 0.01 ± 0.02
sample
glutamine glycine leucine lysine tryptophan tyrosine
response 0.55 5.08 1.59 0.35 2.77 0.17
± · ± · ± ±
0.02 0.05 0.04 0.02 0.20 0.02
“All analyte concentrations are 10 µ , the system shown in Figure 1 was used with the heated reactor at 85 °C. Reagent composition and flow rates are as described in the Experimental Section.
better than 20 nM. This corresponds to less than 300 fmol of NH4+ in the injected sample. This is about a factor of 5 better than the best results we have been able to obtain with the ME reaction system. Reproducibility at levels 1 µ and below is 3.9% mean relative standard deviation and improves at higher levels to 2.7% mean relative standard deviation. The variability is somewhat higher than typical flow-injection results and is in part traceable to the use of an injection valve where a significant fraction of the total volume injected is comprised of valve internal volume. A second source of imprecision is irreproducible self-segmentation induced by bubbles produced in the heated reactor (21). With the heated reactor maintained at 85 °C, the system exhibits linear response behavior in the range 0.25-20 µ NH4+ with a linear correlation coefficient better than 0.999. The calibration shows some curvature at levels below 250 nM (linear correlation coefficient for 50-300 nM is 0.954); however, despite preparation of standards in ammonia-free enclosures and immediate use, contamination from atmospheric ammonia is extremely difficult to eliminate. The problems of external contamination from NH3 are well-known (26), a major source in the laboratory is frequently the human operator—NH3 concentration in expired air is often several hundred parts per billion by volume (27). It is not possible therefore to conclude whether the nonlinearity at the low end is real or a contamination-induced artifact. Above 20 µ , the response shows negative deviations from linearity. If the heated reactor is omitted altogether, less product is formed and the response to 1-1000 µ NH4+ shows excellent linearity (r > 0.999); of course, the limit of detection (LOD) increases. When conducted at room temperature (ca. 22 °C), nonlinearity does not become evident until an NH3 concentration of 2 mM. The throughput rate for 1% carryover between successive samples is 25 samples per hour. In this work, we do not claim to have attained the ultimate in terms of detectability that is possible by this chemistry. The D value in the reaction system (ratio of the signal height in a system with an infinitely long sample loop to that obtained in the real system) was measured, by continuously aspirating a sample in lieu of the carrier, to be 5.0. The dilution of the sample can be much reduced by reducing the OPA and buffered sulfite reagent flow rates (with a corresponding increase in their concentrations), reducing the diameter of the heated reactor (while maintaining the same residence time), =
3, is
and modestly increasing the sample volume. Overall, an order or magnitude improvement in LOD appears feasible but is likely to be experimentally realized only if the entire system is operated in an NH3-free glovebox. A number of environmental water samples have been measured by this technique. Tap water in Lubbock, TX, was found to typically contain 13-15 µ , precipitation 5-9 µ (as a comparison, Hendry and Brezonik (28) report a mean value of 6.7 µ at Gainesville, FL), and water from local playa
1989
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NH/.
lakes ~1 mM
Table I. Relative Response of the Reaction System to Selected Amino Acids0
1,
Relative Response to Amino Acids. Although the OPA-ME-NH3 reaction is slower at neutral pH, Goyal et al.
(20) selected these conditions for the determination of NH3, because prior work (14) established that under these conditions the method is more selective for NH3, relative to amino acids. The responses of the experimental system described to selected amino acids were measured relative to ammonium at a concentration level of 10 µ ; the results are listed in Table I. The method is obviously more selective for NH3 than for amino acids. During the progress of this work, Roach and Harmony (29) and de Montigny et al. (30) reported a new derivatization procedure for amino acids using the naphthalene analog of OPA and cyanide as the nucleophile. Greater stabilities than
the OPA-ME-derivative and better fluorescence detectabilities were reported. Of particular interest to us is that the use of sulfite as nucleophile was also briefly explored. The results for alanine showed more than an order of magnitude decrease in fluorescence intensity in going from ME to sulfite, concordant with our results. Additionally, we found that using cyanide (pH 9.5) instead of sulfite results in a system that is far more sensitive for amino acids than for NH3; this is similar to the behavior of ME at alkaline pH (6, 31). Due to the adverse selectivity and toxicity considerations, further use of cyanide was not pursued. However, we examined the naphthalene analog of OPA for its response to ammonia in the sulfite reaction system. The results were not attractive.
ACKNOWLEDGMENT We thank John N. Marx and William S. Edgemond of this institution for the synthesis of naphthalenedicarboxyaldehyde.
LITERATURE CITED (1) National Academy of Sciences Ammonia; University Park Press: Baltimore, MD, 1979. (2) Tanner, R. L.; Marlow, W. H.; Newman, L. Environ. Sci. Technol. 1979, 13, 75-78. (3) Poirot, R. L; Wlshlnskl, P. R. Atmos. Environ. 1989, 20, 1457-1469. (4) Stevens, R. K.; Dzubay, T. G.; Shaw, R. W., Jr.; McClenny, W. A.; Lewis, C. W.; Wilson, E. W. Environ. Sci. Technol. 1980, 14,
1491-1498. (5) Standard Methods for Examination of Water and Wastewater, 16th ed.; American Public Health Association: Washington, DC, 1985. (6) Roth, M. Anal. Chem. 1971, 43, 880-882. (7) Simons, S. S., Jr.; Johnson, D. F. J. Am. Chem. Soc. 1976, 98,
7098-7099.
(8) Simons, S. S., Jr.; Johnson, D. F. J. Org. Chem.
2886-2891.
1978, 43,
(9) Skoog, D. A. Principles of Instrumental Analysis, 3rd ed.; Saunders: New York, 1984; pp 812-814. (10) Nakamura, H.; Tamura, Z. Anal. Chem. 1981, 53, 2190-2193. (11) Mopper, K.; Delmas, D. Anal. Chem. 1984, 56, 2557-2560. (12) Jacobs, W. A. J. Chromatogr. 1987, 392, 435-441. (13) Saltzman, E. S„ University of Miami, personal communication, 1987. (14) Taylor, S.; Ninjoor, V.; Dowd, D. M.; Tappel, A. L. Anal. Biochem.
1974, 60, (15) Abbas, R.; (16) Danielson, (17) Aokl, T.;
153-162.
Tanner, R. L. Atmos. Environ. 1984, 15, 277-281. N. D.; Conroy, C. M. Talanta 1982, 29, 401-404.
Uemura, S.; Meunemorl, M. Anal. Chem. 1983, 55,
1620-1622.
(18) Riós, A.; Luque de Castro, M. D.; Valcárcel, M. Anal. Chim. Acta 1986, 187, 139-145. (19) Rapsomanikis, S.; Wake, M.; Kitto, A.-M. N.; Harrison, R. M. Environ. Sci. Technol. 1988, 22, 948-952. (20) Goyal, S. S.; Rains, W. W.; Huffaker, R. C. Anal. Chem. 1988, 60,
175-179.
(21) Dong, S.; Dasgupta, P. K. Environ. Sci. Technol. 1987, 21, 581-588. (22) Dasgupta, P. K.; Decesare, K.; Ullrey, J. C. Anal. Chem. 1980, 52,
1912-1922.
(23) Hoffmann, M. R.; Edwards, J. O. J.
2096-2098.
Phys. Chem. 1975,
79,
(24) Takadate, A.; Fujino, H.; Obasa, M.; Goya, S. Chem. Pharm. Bull. 1988, 34, 1172-1175. (25) American Chemical Society Committee on Environment Improvement Anal. Chem. 1980, 52, 2242-2249. (26) Salgne, C.; Klrchner, S.; Legrand, M. Anal. Chlm. Acta 1987, 203,
11-21.
(27) Larson, T. V.; Covert, D. S.; Frank, R.; Charlson, R. J. Science 1977, 197, 161-163. (28) Hendry, C. D.; Brezonik, P. L. Environ. Sci. Technol. 1980, 14,
843-849.
(29) Roach, M. C.; Harmony, M. D. Anal. Chem. 1987, 59, 411-415.
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412
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(30) de Montlgny, P.; Stobbaugh, J. F.; Givens, R. S.; Carlson, R. G.; Srinl·· vasachar, K.; Sternson, L. A.; Hlguchi, T. Anal. Chem. 1987, (31) Llndroth, P.; Mopper, K. Anal. Chem. 1979, 51, 1667-1674.
RECEIVED
This work
was
supported partially by the Electric
Power Research Institute through RP 1630-55 and by the U.S. Department of Energy through DE-FG05-84ER-13281. The contents of this article, have not, however been reviewed by the DOE and no official endorsement should be inferred.
1096-1101.
for review September 27,1988. Accepted December
Simultaneous Quantitative Determination of the Distribution of Dopants in Silicon by High Mass Resolution Secondary Ion Mass Spectrometry Gerhard Stingeder,* Kurt Piplits, Stefan Gara, and Manfred Grasserbauer Institute of Analytical Chemistry, Technical University Vienna, Getreidemarkt 9/151, A-1060 Vienna, Austria
Matthias Budil and Hans Pótzl1 Institute of General Electrical Engineering and Electronics, Technical University Vienna, Vienna, Austria
mechanism. The ratios of phosphorus/marker concentrations
For the study of the diffusion of phosphorus In silicon by mutual diffusion with marker elements (B, As, Sb) It Is necessary to determine the ratios of phosphorus/marker concentrations versus depth with high accuracy. A measurement technique for simultaneous distribution analysis with high mass resolution secondary Ion mass spectrometry has been developed. The signals of SI", P", BSI", AsSi", and SbSi" are utilized for pro4500 Is applied. filing, and a mass resolution of M/AM The precise and reproducible setting of the magnetic field of the analyzer Is controlled In every measurement cycle. The settling time Is determined empirically. Drifts of the analyzer are compensated by transferring the absolute shift of the magnetic field of a reference mass (e.g. 30SI") to the analytical masses. For shallow profiles and low concentrations 4500 and of the markers, measurements with M/AM Mí AM 300 are combined to obtain high sensitivity and depth Information. The detection limits are 1 X 1015 atom cm"3 for B and P, 1 X 1014 atom cm"3 for As, and 4 X 1013 atom cm"3 for Sb. The accuracy Is typically ±25% relative.
depth are one of the input parameters for modeling. Often steep gradients occur; thus the distributions of the dopants should be determined simultaneously to avoid errors in the depth scale owing to consecutive measurements.
versus
EXPERIMENTAL
~
~
~
~
INTRODUCTION In very large scale integration (VLSI) technology process modeling has become an essential development tool (1). The diffusion of B, As, and Sb can be simulated with adequate reliability. The diffusion of P is a very complex process and results at high concentrations in profiles that feature so-called “plateau, kink, and tail” (2) (see Figure 1). Thus models for P have to be established on a physical base, taking into account the diffusion mechanism via point defects in the silicon single crystal (3). It is well-known that Sb diffuses mainly via vacancies and B exhibits an interstitialcy mechanism. For As both mechanisms are equally important. Furthermore, the dopants have different charges when they are incorporated substitutionally in the Si lattice (B“, As"1", Sb+). This means that they can generate different electrical fields, and their diffusion is influenced in a different way by electrical fields. For these reasons during mutual diffusion with P, B, As, and Sb can be used as internal markers to study the diffusion 1
Also Ludwig-Boltzmann-Institute for Solid State Physics, Vien-
na, Austria.
0003-2700/89/0361-0412$01.50/0
SECTION
Secondary ion mass spectrometry (SIMS) measurements were performed with a Cameca IMS 3f instrument using 14.5-keV Cs+ primary ions and detection of negative secondary ions. The signals of the silicon-containing ion species of the dopants are about 1 order of magnitude higher than of the atomic ions. SiP" cannot be utilized, however, because of the interfering 29Si30Si", which corresponds to a background of 3 X 1019 atom cm'3 P and can be separated only with a mass resolution of more than 130000. Thus the signals of Si", P", BSi", AsSi", and SbSi" are detected, and a mass resolution of approximately 4500 is applied to separate the interfering "SPH". Simultaneous Measurement. All signals have to be detected 1 X 106 by the electron multiplier, which can be used up to counts/s. The ion species have to be selected to obtain the maximum dynamic range and sensitivity. If the count rates of 30Si" and 31P~ are too high owing to high erosion rates or P concentrations, their intensities are reduced by detecting only ions with high initial energy (e.g. 50-100 eV). For the molecular ions this is not suitable, because of the steep gradients of the energy distributions at high energies. Thus the lower abundant "Si isotope-containing molecular ion (e.g. 75As"Si") is detected. The main problem of simultaneous measurement with high mass resolution (HMR) is the precise and reproducible setting of the magnetic field. Hysteresis effects are eliminated by cycling many times (~ 20-100) through the measurement sequence. The adjustment of the mass scale is performed by computer control using the centroid algorithm (4, 5). The settling time to obtain a stable signal depends on the mass difference and on the mass resolution and is determined empirically for each combination of the four analytical ions. Typical values are as follows: for switching from mass 30 to mass 31,1.5 s; from mass 31 to 39, 2.5 s; from mass 30 or 39 to mass 103, 6 s; from mass 30 or 39 to mass 149, 10 s. These long waiting times, in addition to integration times of 3-10 s for each element, limit the maximum usable erosion rate and thus the depth resolution. Drifts of the magnetic field setting are caused mainly by temperature changes (air conditioning and warming up of the instrument). To compensate for drifts we have developed a “peak switching autocontrol” (PSAC) computer routine (4, 5). During depth profiling, the magnetic field for each secondary ion can be recalibrated in every measurement cycle. This approach is limited ©
1989 American Chemical Society
