Anal. Chem. 1986, 58,2839-2844
bility could be avoided, and any lasing region within the tuning capability of the optics could be selected. Finally, we should point out that advances in technology could be incorporated to yield a considerably more sensitive system. In the first place, Nd:YAG pumped dye lasers using side pumping and an amplifier stage yield pulses with 100 times greater intensity and stability in this spectral range. In the second place, proximity focused intensifiers are available with much better UV sensitivity. In addition, these devices can be gated to discriminate against Raman photons (30,31). Moreover, these new intensifiers are equipped with phosphors that exhibit linear decay behavior. With all of these improvements, one could anticipate considerable sensitivity improvements, especially in the ultraviolet. But even with the current LVF in its rudimentary state of development, the advantages of multiwavelength excitation coupled with multichannel detection for liquid chromatographic analysis are clear.
ACKNOWLEDGMENT The authors thank David Kalman of the Department of Environmental Health a t the University of Washington for kindly supplying polyaromatic hydrocarbon reference materials and for the soil extract sample. The authors also thank David H. Burns and Barry V. Pepich for useful discussion and criticism. Registry No. BBF,205-99-2; PER, 198-55-0; BKF,207-08-9; BAP, 50-32-8; 3-OH-BAP, 13345-21-6;FL, 86-73-7;AC, 83-32-9; PH, 85-01-8; ANT, 120-12-7;FLU,206-44-0; PY, 129-00-0;CHR, 218-01-9; BAA, 56-55-3; D(a,h)A, 53-70-3; BGP, 191-24-2; IP, 193-39-5; 9-MEA, 779-02-2. LITERATURE CITED Richardson, Jeffery H.; Ando, M. E. Anal. Chem. 1977, 49, 955-959. Weeks, Stephan J.; Haraguchi, Hiroki; Winefordner, James D. Anal. Chem. 1979, 50, 360-368. Ishibashl, Nobuhiko; Ogawa, Teiichiro; Imasaka, Totaro; Kunitake, Mikio Anal. Chem. 1979, 57,2096-2099. Strojny, Norman; de Silva, Arthur F. Anal. Chem. 1980, 52, 1554- 1559. Voigtman, Edward G.; Jurgensen, Arthur; Winefordner, James D. Anal. Chem. 1981, 53, 1921-1923. Furuta, Naoki; Otsuki, Akira Anal. Chem. 1983, 55, 2407-2413. Hirschfeid, Tomas Anal. Chem. 1985, 52, 297A-312A. Hershberger, Leon W.; Callis, James B.; Christian, Gary D. Anal. Chem. 1979, 57, 1444-1446.
2839
Hershberger. Leon W.; Callis, James B.; Christian, Gary D. Anal. Chem. 1981, 53, 971-975. Fogarty, Michael P.; Shelly, Dennis C.; Warner, Isiah M. HRCCC, J . High Resolut . Chromatogr . Chromatogr . Commun . 1981, 4 , 561-568. Ho, Chu-Ngi; Christian, Gary D.; Davidson, Ernest R. Anal. Chem. 1978, 50, 1108. Ho, Chu-Ngi; Christian, Gary D.; Davidson, Ernest R. Anal. Chem. 1980, 52,1071. Ho, Chu-Ngi; Christian, Gary D.; Davidson, Ernest R. Anal. Chem. 1982. 5 4 . 1291. T a l i : Yair; Baker, Donald C.; Jadamec, James R.; Saner, William A. Anal. Chem. 1978, 5 0 , 936A-952A. Shectman, Stephen A.; Hiltner, Wiiiiam A. Pub/ Astron. SOC. Pac. 1976, 88, 960-965. Davis, Marc; Latham, David W. Proc. SPIE-Int. SOC. Opt. Eng. 1979, 772, 71-81. Ryan, Mary A.; Miiier, Robert J.: Ingle, James D., Jr. Anal. Chem. 1978, 5 0 , 1772-1777. Johnson, Craig, R.; Asher, Sanford A. Anal. Chem. 1984, 5 6 , 2258-2261. Gluckman, Jennifer C.; Shelly, Dennis C.; Novotny, Milos V. Anal. Chem. 1985, 5 7 , 1546-1552. Skoropinski,D. Bruce Ph.D. dissertation, University of Washington, Seattle, WA, 1985. Warner, Isiah M.; Patonay, Gabor; Thomas, Mark P. Anal. Chem. 1985, 57, 463A-483A. Roncin, Jean-Yves; Damany, Henri Rev. Sci. Instrum. 1981, 52, 1922-1 923. Lawler, James E.; Fitzsimmons, William A,; Anderson, Lawrence W. Appl. Opt. 1976, 75, 1063-1090. Talmi, Yair; Busch, Kenneth W. I n Multichannel Image Detectors; Talmi, Y., Ed.; American Chemical Society: Washington, DC, 1983; ACS Symposium Series, No. 236, Chapter 1. Stowe, Grady, Varo Electron Devices, Garland TX, private communication. Jerina, Donald M.; Yagi, Hiroto; Hernandez, Otheiio; Dansette, Pamela M.; Wood, Alien W.; Levin, William; Chang, Robert L.; Wisiocki, Peter G.; Conney, Allen H. I n Carcinogenesis, Vol. 7 . Polynuclear Aromatic Hydrocarbons: Chemistry, Metabolism and Carcinogenesis ; Freudenthai, Ralph I., Jones, Peter W., Eds.; Raven Press: New York, 1975; pp 91-1 13. Cole, Thomas; Riggin, Robert; Giaser, James I n Polynuclear Aromatic Hydrocarbons, 5th International Symposium, 7980 ; Cooke, Marcus, Dennis, Anthony J., Eds.; Battelle Press: Columbus, OH, 1981; pp 439-454. Ogan, Kenneth; Katz, Eiena, Slavin, Walter Anal. Chem. 1979, 5 7 , 1315-1320. Nielsen, Torben J. Chromatography 1979, 770, 147-156. King, Nathan S. P.; Yates, Gerald J.; Jaramiilo, Samuel A,; Ogle, John W.; Deutch, James L., Jr. Proc. SPIE-Int. SOC. Opt. Eng. 1981, 288, 426-433. Lieber, Albert J. Rev. Sci. Instrum. 1972, 4 3 , 104-108.
RECEIVED for review February 28, 1986. Resubmitted July 10, 1986. Accepted July 10, 1986. This work was supported in part by NIH Grant No. GM 22311.
Trace Determination of Aqueous Sulfite, Sulfide, and Methanethiol by Fluorometric Flow Injection Analysis Purnendu K. Dasgupta* and Huey-Chin Yang Department of Chemistry and Biochemistry, Texas Tech Uniuersity, Lubbock, Texas 79409-4260 Preservationof sulfRe, sulfide, and methanethiol In buffered formaldehyde and oxaklihydroxamlc acld stabilizers has been studled. Flow InJectlonanalysis procedures that involve T mixing or membrane-based reagent Introduction have been developed for the fast (24 sampieslh) analysis of these anions based upon the reaction wRh N-acrldlnylmaieimldein a water-N,N4methylformamlde medium to form a fluorescent product. Detection llmlts are 0.04, 0.60, and 0.80 pM, respectlvely, for the three sulfur species; dlfferentlai analysls Is possible.
Trace determination of reduced sulfur anions in aqueous
solution is important in atmospheric analysis, especially in the elucidation of various aspects of the occurrence, distribution, and chemistry of atmospheric sulfur compounds as well as in understanding the phenomenology of acid precipitation (1,2). Atmospheric sulfur gases such as SO2, H2S,and CHBSHare often determined after collection in an aqueous absorber (3). Consequently, improving detection limits in the determination of the corresponding anions in aqueous solution ultimately benefit the gas-phase detection limits as well. The advent of the diffusion scrubber (4-6) has accentuated the need for fast, sensitive aqueous phase continuous flow analytical procedures; these are ideally coupled to the scrubbers. We are interested in fast and sensitive flow injection analysis (FIA)procedures for this reason, and direct application of such
0003-2700/86/0356-2839$01.50/00 1986 American Chemical Society
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ANALYTICAL CHEMISTRY, VOL. 58, NO. 13, NOVEMBER 1986
procedures to determine trace quantities of atmospheric constituents such as S(1V) or H 2 0 2in cloudwater or precipitation is also of interest (7, 8). The collection of atmospheric sulfur dioxide in tetrachloromercurate(I1) and its subsequent colorimetric determination by reaction with formaldehyde and pararosaniline, due originally to West and Gaeke (9),is likely still the most widely used wet chemical for atmospheric SO,; in modified form, it is the basis of the federal reference method (10). Dilute buffered formaldehyde solution was introduced as an absorber for SO2collection in 1980 (11)and it has been shown that the thermal and photochemical stability of the SO, collected as hydroxymethanesulfonate in such a medium is substantially superior to that of the mercury complex formed in the West-Gaeke method (12). More recent measurements (13-15) of the formaldehyde-bisulfite-hydroxymethanesulfonate equilibrium system indicate that the formation constant of the addition compound is in fact 2 orders of magnitude higher than that previously reported (11). The formaldehyde absorber procedure has been used successfully for routine monitoring applications (16, 17) and has been automated for atmospheric analysis by segmented continuous flow analysis (18) or nonsegmented membrane-based FIA (7) methods. The chemistry of the Schiff reaction, a singularly important reaction in histochemistry (19),involving a ternary reaction of pararosaniline, formaldehyde, and S(IV) in various temporal combinations, has been elucidated (11,20, 21). It is understood to be an "indicator pK-shift reaction" (22)that can successfully, and with the benefit of greater sensitivity, fluorescent acid-base inbe extended to nonfluorescent dicators such as 1-aminonaphthalene (22)or anthranilic acid (23)compared to the colorless-colored transition that occurs with pararosaniline (11). Unfortunately, the kinetics of the reactions involved are such that whenever S(1V) stabilized in a formaldehyde solution is reacted, after base-induced decomposition, with an acidic solution, the analytical response decreases nonlinearly at the low end of the analyte concentration scale (7.18,22,23). Additionally, the calibration slopes in the automated versions, in our experience, can be acutely dependent on mixing conditions (24). Further, the reaction is slow relative to optimal FIA time scales. An analytical reaction that can be carried out in an alkaline medium may avoid some of these difficulties and be compatible with the formaldehyde absorber; alternatively, a different sulfite stabilizer, e.g., oxaldihydroxamic acid (ODHA) may be sought (25,26). Maleimide-based fluorogenic probes for thiols, sulfides, and sulfites are well-known (27). One of the recent entries among these is nonfluorescent Nacridinylmaleimide (NAM),which reacts with sulfites, sulfides, and thiols under alkaline condition (28-36) to form 3sulfonato- or 3-mercapto-NAM. Selective determination of sulfite is possible in the presence of metal ions such as divalent Co, Hg, or Cu (29). The recommended reaction conditions (1 h , 35 "C), however, preclude facile FIA adaptation. We report here membrane-based or conventional FIA procedures that integrate the use of the formaldehyde or ODHA stabilizers with a modified room temperature NAM reaction for the differential determinations of sulfite, sulfide, and methanethiol. Stabilities of such species in dilute solutions in formaldehyde or ODHA stabilizers are also reported.
-
EXPERIMENTAL SECTION Reagents and Equipment. ODHA was synthesized according to the method of Paul and Gupta (25)and NAM was synthesized by the method of Nara and Tsuzimura (28) or obtained from Dojindo Laboratories (Tokyo, Japan) or Sigma Chemical (St. Louis, MO). All other reagents used were of analytical reagent grade. The fluorescence derivatization reagent is prepared by dissolving 1 mg of NAM in 20 mL of NJV-dimethylformamide (DMF)
Stabilizer
- Injection
Carbonah Carrier Buffer 0.044 NAM
a.-ad\
-''\
Fluorescence Detector
J!-~--.
,044
U
Figure 1. Basic FIA system in the conventional T-mixing mode. The lengths represented by a, b, and c are 24 cm, 28 cm, and 125 cm, respectively, 0.8 m m i.d. tubing. D value for a 225-,uL sample is 1.6.
Pump mt.in'
n
Knotted Line Air
\ FJ
Pressure
Flgure 2. Membrane-based FIA system. I n the presence of HCHO, additional flowstream of KCN is used. See text for details. a, b,
an
c, and d, respectively, are 24 cm, 15 c m (including reactor), 39 c m (including reactor), and 148 cm.
followed by 20 mL of water. The solution is not stable for extended periods, storage at -5 "C in the dark is recommended. The formaldehyde absorber compostion was 7 mM HCHO and 1mM potassium acid phthalate, pH 4.0 (11). The ODHA absorber composition was 5 mM ODHA and 1 mM potassium acid phthalate, pH 4.0. Each absorber may be modified for sulfite selectivity by the addition of 0.5 mM CdCl,. The carbonate-bicarbonate buffer is made by dissolving -2.3 g of KZCO, in 250 mL of a 0.5 M solution of KHC03 to obtain a pH of 9.2. Note that a mixed NAM reagent-buffer solution is too unstable to be conveniently used. Standard analyte solutions absorber media were prepared from NaHSO,, Na2S, liquid CH,SH, (CH3)2S,and (CH3)2Sz. The stabilities of the reduced sulfur species in the aqueous absorbers were measured under three storage conditions: at ambient laboratory temperature and exposed to light, at ambient temperature in the dark, and under refrigeration in the dark. The decay measurements were made by manual analogs of the FIA procedure described below with a reaction time of 5 min on a Perkin-Elmer LS-5 instrument under optimal excitation conditions (Aex 360 nm, 15-nm slit; ,A, 435 nm, 20-nm slit). Flow Injection Analysis Systems. The experimental arrangements involved either conventional tee mixing (Figure 1) or membrane-based reagent introduction (Figure 2). In Figure 1,the carrier:carbonate buffer:NAM flow rate ratio is 1Ol:l (carrier flow rate 440 pL/min) and the reaction time (after NAM introduction) is 50 s. With the sample volume of 225 pL (0.8 X 450 mm loop) the D value (37) at the detector was 1.6. The injection valve used was either an electrically actuated (Type HVLX 6-6, Hamilton Co., Reno, NV) or a manual (Type 50, Rheodyne, Inc., Cotati, CA) six-portvalve. A peristaltic pump (Minipuls 2, Gilson Medical Electronics,Middleton, WI) was used for pumping. The detector used was a Fluoromonitor I11 (LDC/Milton Roy, Riviera Beach, FL) equipped with a Cd-line source (326 nm), and a high pass filter (50% cutoff point 370 nm) on the emission side. Unless otherwise indicated, all tubing was 0.8 mm i.d. poly(tetrafluor0ethylene) tubing (20 SW, Zeus Industrial Products, Raritan, NJ). In the membrane-based system, the carrier was pumped at a rate of 440 pL/min through the same valve and sample loop as above into a passive membrane reactor (0.8 mm diameter poly(tetrafluorethylene)filament-filled 20 mm long length of tubing made of Nafion 811x perfluorinated membrane, Perma-Pure Products, Toms River, NJ; for connection details, see ref 8) immersed in a solution of concentrated ",OH for ammonia in-
ANALYTICAL CHEMISTRY, VOL. 58, NO. 13, NOVEMBER 1986
2841
.I r...
I
340 310-
300.
"t
40
60
/I:: 10,
d
40
30
* O t
I 0
/ I 5
I
I
10
15
I
20
Tirne(days) Figure 3. Efficacy of different stabilizers for reduced sulfur anions: (a) sulfite in 5 mM ODHA-1 mM KHP-0.5 mM CdCI,; (b) CH,SH in 7 mM HCHO-1 mM KHP; (c) CH3SH in 2.5 mM ODHA-1 mM KHP; (d) sulfide in 7 mM HCHO-1 mM KHP. Percent of analyte remaining, on a logarithmic scale, is plotted against the time of storage. The storage regimes are refrigeration (closed symbols), ambient temperature and dark (half-closed symbols), and ambient light and temperature (open symbols). Starting concentrations of sulfur analytes in each case is 1.9-2.0 KM.
-
troduction to raise the pH to 10 and then through a pressurized porous membrane reactor (0.8 mm diameter TFE filament filled porous polysulfone membrane tube, 20 mm long, 0.1 pm mean pore size, 1 mm i.d., A/G Technology, Needham, MA; for connection details, see ref 8) immersed in a solution of the NAM reagent for the introduction of the same. A superincumbent air pressure of 11.5 psi was found adequate to obtain the optimal NAM introduction rate, equivalent to -44 pL/min of conventional addition. The same reaction time, 50 s, was used. The D value for a 225 p L sample was 1.8. Formaldehyde-stabilized sulfite solutions can be assayed by the NAM reaction only if the formaldehyde is sequestered, e.g., with cyanide (vide infra). Both types of FIA systems above were modified by the addition of a 22 pL/min pumped channel of 0.3 M KCN which was tee-mixed, after the sample introduction valves, with the carrier, prior to the introduction of buffer or NAM. This is shown as the dotted pumping channel in Figure 2. The closed nature of the FIA system permits safe handling of cyanide. The waste is collected into a container containing alkaline hypochlorite (Clorox). The introduction of cyanide through a membrane reactor is possible but was not investigated.
RESULTS AND DISCUSSIONS Stability of Sulfite, Sulfide, and Methanethiol in Buffered Formaldehyde and ODHA Absorbers. Sulfite. Sulfite stability in buffered formaldehyde has been extensively studied in the past (12) and is highly stable with a first-order decay constant of 6.51 x day-' at 22 "C and 1.97 x day-' at 50 "C. Sulfite in ODHA solution is acceptably stable over week-long periods only if stored refrigerated at pH 4 (Figure 3a). The unbuffered absorber described in ref 25
,
,
,
,
,
l
,
,
1
1
.
1
l
l
l
l
l
'
,
I
I
1
,
leads to unacceptably high decay rates, especially when exposed to ambient light. Malonyl and succinyl analogues of ODHA were synthesized by a procedure similar to that described for ODHA (%) but were found to be inferior to ODHA in stabilizing S(1V). The addition of 0.5 mM CdClz to either the formaldehyde or the ODHA absorber does not significantly influence the decay rate. The first-order constant for the decay of sulfite in units of inverse days in the ODHA-KHPa t 5 "C and CdClz absorber shown in Figure 3a is 3.5 X a t 22 "C both under dark storage and 1.8 x 1.1x a t ambient light and temperature (-22 "C). Methanethiol. Methanethiol is substantially more stable in the formaldehyde absorber (Figure 3b) compared to the ODHA absorber (Figure 3c). The respective first-order decay constants under the three storage regimes specified previously 5.7 X and 2.4 X and 2.2 X 6.2 are 2.3 x X 6.8 X day-' for the two respective absorbers. Refrigerated storage in the HCHO absorber is acceptable over week-long periods. Sulfide. Sulfide is not adequately preserved by either absorber (Figure 3d). The first-order constants for the decay of sulfide in the formaldehyde absorber, the better of the two, under the three storage regimes (dark refrigerated, dark and ambient temperature, ambient light and temperature) are 6.9 X 8.8 X and 1.4 X lo-' day-', respectively. Temporal and Signal Optimization. The sulfite assay procedure with the NAM reagent as given by Meguro and Takahashi (36) recommends an 1-h reaction time a t 35 "C; this is not easily adapted to FIA systems. The effect of the reaction pH (apparent pH) is also significant: In manual assays for sulfite following the published procedure (36), optimal results are obtained only within pH 8.6-9.2; response is very poor at pH 5 8.0 or I 9.8. NAM is virtually insoluble in pure water. Due to its high volatility, we sought to replace acetone used as solvent in previous NAM procedures (28, 36) by some other solvent. (Note that the introduction of neat organic solvents presents no special problem with the porous TFE membrane based pressurized reactor; this is not the case for common peristaltic pump tubes). During these studies, it was discovered that the use of dimethyl sulfoxide (Me2SO)or DMF as solvent not only markedly increases the fluorescence signal but increases the reaction rate as well (Figure 4). However, NAM is rather unstable in neat DMF or MezSOeven when stored refrigerated in darkness. The stability of NAM solutions in 5050 DMF:water was found to be acceptable (>3 days stored refrigerated). It was also discovered that the DMF concentration necessary for the optimal rate of fluorescence enhancement is small, large amounts actually result in lower enhancement
2842
ANALYTICAL CHEMISTRY, VOL. 58,NO. 13, NOVEMBER 1986
Table I. Summary of Analytical Results sensitivity, mV/wM (LOD, pM) S2CHBSH
so:-
carrier A. ODHA-KHP"
134.03 (0.0806)
B. A + 0.5 mM Cd2+ C. HCHO-KHP D. C + 0.5 mM Cd'+ E. HCHO-KHP, CN- modified' F. ODHA-KHP, CN- modifiedC
as in A
9.34 X lo-* (26)" as in C 80.13 (0.0374) e
10 (0.60) -0 -0 -0 15.03 (0.70) e
4.48 (2.0)
-0 6.07* (0.8) -0 3.9 (1.54) e
(CHdzS 1.14 X lo-' (5.30) as in A ndd nd nd nd
Single pumped channel, membrane reactor system. Two pumped channels, membrane reactor a System with three pumped channels. system. dnd, not determined. eSensitivityessentially as in A, LOD's are somewhat worse due to increased blank values. 1
F.I.1
1
I
80 /
l
l
l
1
2
3 4
,
l
1
l
l
,
l
l
l
,
5 6 7 8 9 101112131415 TIME( m i n )
Figure 5. Temporal fluorescence development for a 2 p M S(IV) sample as in Figure 4, as a function of NAM concentration and DMF content of the reagent: 0,0.002 w/v % NAM in 33 vol YO DMF; V, 0.002 w/v YO NAM in 50 vol YO DMF; A,0.0025 w/v % NAM in 50 vol % DMF; 0, 0.004 w/v % NAM in neat DMF; 0 , 0.005 w/v % NAM in neat DMF.
factors. Figure E shows the temporal development of the fluorescence signal as a function of NAM concentration and DMF concentration in the NAM solution. For optimal results, the procedure described in this paper uses a 0.0025 (w/v) NAM solution in 1:l (v/v) DMF:water. We examined the utility of micellar (hexadecyltrimethylammonium chloride and Triton-X-100) solubilization and potential catalysis and fluorescence enhancement; results were not attractive. System Performance. Sulfite. With the ODHA absorber with or without Cd2+,excellent results were obtained with either the T-based or membrane-based system. Data are indicated in Table I for the T-based system; comparable results were obtained with the membrane based system with 3.0 M NHIOH exterior to the passive membrane reactor, yielding an effluent pH of 9.5. However, this is a relatively low blank method. With the low-pulsation pumping system used, no significant difference between T-based and membrane-based reagent introduction system is expected. A typical calibration plot for the T-based system is shown in Figure 6a. Because of the exceptional stability of sulfite in formaldehyde solutions, we wished to apply the NAM reaction to formaldehyde stabilized S(1V) samples. A special effort was necessary since it was discovered that essentially no signals are obtained for sulfite samples preserved in the formaldehyde stabilizer. Based on the present values for the equilibrium constant of the formaldehyde-bisulfite-hydroxymethanesulfonate equilibrium (13-151, some, but not quantitative, interference may be expected for micromolar levels of sulfite
(a) Time Flgure 6. Calibration plots: (a) sulfite in three pumped channel system, ODHAstabilizer(1,3.12pM;2,2.81;3,2.50;4,2.18;5,1.87;6, 1.56; 7, 1.25; 5, 0.94;9, 0.62; 10, 0.32); (b) CHBSH in single pumped channel, membrane reactor system, HCHO stabilizer (1, 20 pM; 2, 10; 3, 5.0; 4, 1.0); (c) sulfide in two pumped channel, HCHO stabilizer, CN--modified during analysis (1, 50 pM; 2, 40;3, 30; 4,20; 5, 10).
in the presence of 7 mM HCHO (absorber composition) at a reaction pH of 9-10. Experimentally, it was found that sulfide behaves in a manner similar to sulfite in the presence of HCHO; however, no significant inhibition of fluorescence development is observed in the case of methanethiol. Unsuccessful efforts were made to sequester formaldehyde with ammonia/hydroxylamine/hydrazine/aniline to allow NAM-sulfonate to be formed. Cyanide, when used in excess, was found to be the only agent that sequestered formaldehyde in the desired manner. Although the blank values with either absorber are much higher (approximately 5 times) in the presence of cyanide (presumably due to the formation of the nitrile derivative of NAM), and the calibration slope (sensitivity) is also somewhat lower (possibly indicating that the formaldehyde effect is not totally eliminated), the membrane-based FIA system permits sufficiently good precision to attain a limit of detection of 37 nM (8 pmol in the sample volume), the best yet attained, to our knowledge, for the
ANALYTICAL CHEMISTRY, VOL. 58, NO. 13, NOVEMBER 1986
determination of sulfite (Table I). It is noteworthy that this limit of detection is attained with an excitation wavelength far from optimal, dictated by available equipment. Methanethiol. The sensitivity of the NAM reaction for methanethiol is 25-30 times lower than that of sulfite on a molar basis; the limit of detection dete?iorates proportionately. Formaldehyde, the preferred stabilizer, actually permits a slightly more sensitive procedure compared to the ODHA preservative. For the membrane-based systems, the optimal ammonia concentration and effluent pH were determined, respectively, to be 17 M and 10.6 for the formaldehyde stabilizer. Sub-micromolar detection limits can be attained (Table I) and a typical calibration plot is shown in Figure 6b. Sulfide. The sensitivity for determining sulfide in manual assays by the NAM reaction under optimal optical conditions and a 5-min reaction time is half that for sulfite under analogous conditions. For inexplicable reasons, the sensitivities obtained in the FIA system were significantly worse. However, sub-micromolar limits of detection are attainable for the ODHA stabilizer (A, Table I) and both cyanide-added procedures (E and F, Table I) in membrane-based FIA systems. A typical calibration plot for sulfide in system E is shown in Figure 6c. Dimethyl Sulfide and Dimethyl Disulfide. Dimethyl disulfide does not react with NAM under any conditions tested by us. Nevertheless,not all disulfides may behave in the same fashion (29). Dimethyl sulfide is detectable only at very high concentrations (detection limit 0.5 mM) with a sensitivity of less than 0.0170of that for sulfite. Normally this small signal could be attributed to impurities present in the commercial product; however, the signal persisted upon cadmium addition and is not due to sulfide. The sample throughput rate for any of the FIA configurations described above was 24 per hour. Selective Determinations: Cadmium Effect. Preliminary studies showed that Cd2+was the most effective among Cd2+,Cu2+,and Hg2+for eliminating the signal due to sulfide and methanethiol and a concentration of 0.5 mM Cd2+was judged adequate for likely sulfide and methanethiol concentrations in atmospheric samples. Much higher concentrations of Cd2+led to precipitation of CdS when the solution is made alkaline; turbidity is also predictably observed at significant concentrations of sulfide, although no problems due to light scattering etc. were encountered. The ODHA absorber therefore permits selective determination of sulfite when cadmium is added. Because the use of the formaldehyde absorber requires that cyanide be added during analysis, and cadmium is expected to be complexed by cyanide, cadmium and any other metal is not expected to completely eliminate the divalent sulfur anion signals and this aspect was therefore not explored. In the absence of cadmium, signals from the three sulfur anions in ODHA absorber were found to be additive. On the other hand, direct selective determination of methanethiol is possible when the formaldehyde absorber is used. If cyanide is added during the analysis procedure, the analytical signal represents the sum of all three sulfur species. Consequently, determinations in systems A (or E), B, and C (Table I) provide complete analytical results for the mixture of the three analytes. While B and C provide direct measures of sulfite and methanethiol, respectively, the sum of all three are represented by A (or E) and sulfide may be obtained by difference. Due to the significant differences in sensitivities of the method(s) to the various analytes, relative accuracy for sulfide is poor if comparable amounts of all anions are present. It should be noted however that no specific optimization of analytical systems A and E were performed with respect to the sulfide signal, rather the conditions were optimized for
1 1
I
200
I
4
1
1
1
I
2843
' 1 & l
Flgure 7. Effect of formaldehyde addition to ODHA-preserved S(IV) samples upon signal height (arbitrary units), no cyanide added (system A, Table I): (A) 3.1 pM S(IV); (B) 1.6 pM S(IV).
Table 11. S(1V) Content in Fogwater: Riverside, CA, March 1985 sample ID
[S(IV)],"r M
sample ID
[S(IV)]: rM
2A 3A 4A
148 f 1.1 78.8 1.0 134 f 0.4
5A 6A
107 f 0.5 107 f 0.5
*
Triplicate determination after dilution. Concentrations are corrected for dilution due to subsequent preservative addition. Equal amounts of double strength ODHA absorber and fogwater sample were mixed before storage. sulfite analysis. It is possible that sulfide sensitivity may be enhanced under different experimental conditions. Interferences: Analysis of Environmental Samples f o r Sulfite. The NAM reaction, like reactions of other fluorogenic maleimides (27) is highly specific for the sulfur anions, However, negative interference from agents such as formaldehyde, as noted previously, may be present for the non-cyanide type analysis systems. The effects of various concentrations of formaldehyde on sulfite analysis in systems A and F were studied. No effeds were found for formaldehyde concentrations up to 3 mM for the cyanide-modified system (F). The results for system A are shown in Figure 7. The formaldehyde effect is particularly important because it can be a significant constituent of atmospheric water, especially in photochemically active urban atmospheres (38-40). Other reactive carbonyl compounds are also usually present (41). While explicit studies with other potential aldehydes were not carried out, it is reasonable to expect that the cyanide addition will effectively sequester them as well. It is not possible however to predict the degree of interference such other compounds may pose without cyanide addition. Fogwater samples collected by a modified version of the collector described in ref 42 were preserved in ODHA (so as to permit measurement of HCHO) and analyzed by system F. In agreement with very high levels of sulfite previously reported for such samples (39,40), the samples required large dilution with blank for convenient analysis. The results are shown in Table 11. CONCLUSIONS We have developed a sensitive and reasonably fast FIA procedure to determine sulfte in the presence of formaldehyde and describe adequate means of preserving sulfite in environmental samples. Under favorable circumstances, simultaneous differential determination of sulfide and methanethiol is possible.
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Anal. Chem. 1986, 58, 2844-2847
ACKNOWLEDGMENT We thank J‘ Munger9 Engineering Science, California Institute of Technology, for collecting and supplying the fogwater samples.
(23) McDowell, W. L.; Dasgupta, P. K. Atmos. Environ. 1984, 18, 2209-2216. (24) McDowell, W. L.; Dasgupta, P. K., unpublished results, Texas Tech University, 1983. (25) Paul, K. R.; Gupta, V. K. Atmos. Environ. 1983, 17, 1773-1777. (26) Dasgupta. P. K. Atmos. Environ. 1984, 18, 477-478. (27) Haugland, R. P. Molecular Probes Handbook of Fluorescence Probes; Molecular Probes: Junction City, OR, 1965. (28) Nara, Y.; Tsuzimura, K. Bunseki Kagaku 1973, 2 2 , 451-452. (29) Takahashi. H.; Nara, Y., Tsuzimura, K. Agric. Biol. Chem. 1978, 4 0 , 2493-2494. (30) Takahashi. H.; Nara, Y., Tsuzimura, K. Agric. Biol. Chem. 1978, 4 2 , 769-774. (31) Machich, M.; Takahashi, T.; Itoh, K.; Sekine, T.; Kanaoka. Y. Chem. Pharm. Bull. 1978, 2 6 , 596-604. (32) Nara, Y.; Tsuzimura, K. Agric. Biol. Chem. 1978, 42, 793-798. (33) Takahashi, H.; Nara, Y.; Meguro, H.; Tsuzimura, K. Agric. Biol. Chem. 1979, 43, 1439-1445. (34) Takahashi, H.; Yoshida. T.; Meguro, H. Bunseki Kagaku 1981, 30, 339-34 1. (35) Takahashi, H.; Nara, Y.; Yoshida, T.; Tsuzimura, K.; Meguro, H. Agric. Biol. Chem. 1981, 45, 79-85. (36) Meguro, H.; Takahashi, C. Anal. Lett. 1983, 16(A20), 1625-1632. (37) Ruzicka, J.; Hansen, E. H. Flow Injection Analysis; Wiley: New York, 1981. (38) Richards, L. W.; Anderson, J. A,; Blumenthal, D. L.; McDonald, J. A,; Kok, G. L.; Lazrus, A. L. Atmos. Environ. 1983, 17, 911-914. (39) Munger, J. W.; Jacob, 0. J.; Hoffmann, M. R. J . Atmos. Chem. 1984, 1 , 335-350. ( 4 0 ) Munger, J. W.; Tiller, C., Hoffmann, M. R. Science 1986, 231, 247-249. (41) Grosjean, D.; Wright, B. Atmos. Environ. 1983, 17, 2093-2096. (42) Jacob, D. J.; Waldman, J. M.; Haghi, M.; Hoffman, M. R.; Flagan, R. C. Rev. Sci. Instrum. 1988, 5 6 , 1291-1293.
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RECEIVED for review April 21, 1986. Accepted July 14, 1986. We gratefully acknowledge the support of the Electric Power Research Institute through RP 1630-28for making this work possible.
Measurement of Electron Diffusion Coefficients through Prussian Blue Electroactive Films Electrodeposited on Interdigitated Array Platinum Electrodes B. J. Feldman’ and Royce W. Murray* Kenan Laboratories
of
Chemistry, University of North Carolina, Chapel Hill, North Carolina 27514
Electron mobility In electroactive polymers, measured as the dlffuslon coefflclent D,, can be measured by deposltlng the polymer over fingers and Insulating gap of an Interdigitated array electrode (IDA) and adjusting the electrode potentials so as to oxidize and reduce the polymer fllm at opposing, adjacent finger electrodes. A parallel plate theory for De and the llmlting current flow through the polymer fllm in the IDA gap Is evaluated with Prusslan Blue as the exemplary electroactlve material. The theory accurately describes experlmental behavior for electrodeposited films thick enough to give unlform coatings within the 2.5-pm gap but thinner than the 2.5-pm gap.
Of special interest is the coating of such electrodes with electroactive films. Wrighton and co-workers have shown that microarray electrodes coated with conducting organic polymers such as poly(pyrro1e) ( I , 2 ) and poly(viny1ferrocene) (5)mimic the behavior of solid-state diodes (5)and transistors (1-4) and can act as chromatographic detectors (4). We have shown (8) that an interdigitated array (IDA) electrode coated with Prussian Blue exhibits understandable voltammetry and electrochromism in gas-phase media. Wohltjen and co-workers (IO, 11) have also described gas phase sensor experiments with solid-state “chemiresistors” constructed by applying Langmuir-Blodgett films of nickel phthalocyanines to IDA’S. Coated microarray electrodes and IDA’S additionally find important applications to fundamental electron transport studies, including conductivity as a function of potential for conducting organic polymers (1-4) and the environmental (solvent, ions) dependence of electron hopping rates in crystalline Prussian Blue (8). Whether an IDA coated with an electroactive polymer is used for practical or for more theoretical purposes, a proper measurement of the electron conductivity of the polymer is
Microlithographically defined microarray (1-5) and interdigitated array (6-21) electrodes have been recently shown to have impressive utility in electrochemistry and in sensors. ‘Present address: IBM Almaden Research Center, 650 Harry Rd., K34/802, San Jose, CA 95120-6099. 0003-2700/86/0358-2844$01.50/0
C
1986 American Chemical Society
