Anal. Chem. 1988, 60, 2791-2796
required by this method that the relative ion signal intensities represent the proportion of phospholipids in the bacteria, only that they be reproducible and additive. A test of this requirement has been our ability to assess the composition of mixtures of bacteria with reasonable accuracy (20). The CNL approach increases specificity for phospholipid classes. It also suggests a protocol for rapid classification and identification of bacteria which would combine the linear regression analysis (based upon mass and intensity in the normal magnetic scan) with a more rational and interpretative approach. The negative ion FAB spectra would be used for library matching, but the positive ion CNL scan for a loss of 155 mass units (for example) would restrict the search to those bacteria that have been determined to contain lipids with a methylphosphoethanolaminehead group. Other approaches might depend less heavily on the normal mass spectra (which reflect growth conditions) and more upon the presence, absence, or relative intensity of specific polar head groups that are ascertained by the CNL spectra. In either case, the increased specificity for biomarkers in the native bacteria (through selective desorption and specific fragmentation) provides the opportunity for developing rapid, computerized identification.
LITERATURE CITED Kates, M. In Advances In UpM Research; Paolenl, R.. Ed.; Academic: New York, 1982; Vol. 2, pp 17-70. Lechevaller, M. P. CRC Crn. Rev. MImb&l. 1977, 109-210. Goldflne, H. In Membrane Lip& of prokeryotes; Razln, S.. Rottem, S., Eds.; Acedemlc: New York, 1982; pp 1-43. Heller, D. N.; Fenselau, C.; Cotter, R. J.; Demlrev, P.; Otthoff, J. K.; Honovlch, J.; Uy, M.; Tanaka, T.; Klshlmoto, Y. Blochem. BIophys. Res. Commun. 1987, 142, 194-199. Heller, D. N.; Cotter, R. J.; Fenselau, C.; Uy, 0. M. Anal. Chem. 1987, 5 9 , 2806-2809. Moss, C. W. J . Chromatog. 1981, 203, 337-347. Pramanlk, B. N.; Zechman, J. M.:Des, P. R.; Bartner, P. L.; Horan, A. C.; Morton, J. B. P m . ASMS Conf. Mass Spectrom. Allied Topics, 34th 1988, 122-123.
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Dasgupta, A.; Ayanoglu, E.; Tomer, K. 8.; Djerassl, C. Chem. Fhys. LipMs 1987,43, 101-111. Chltton, F. H.. 111; Murphy, R. C. Blamed. Envkon. Mass Spectrom. 1986, 13, 71-76. Kim, H.-Y.; Salem, N., Jr. Anal. Chem. 1987, 5 9 , 722-728. Heller, D. N.; Fenselau, C.; Cotter,R. J.; Demlrev, P.; Olthoff, J.; Honovlch, J. Roc. Annu. Conf. Mess Specirom. ANM Topics, 34th Clnclnnati OH (1986) pp 872-873. Munster, H.; Stein, J.; Budzlklewlcz, H. Blamed. Envkon. Mass Specfrom. 1986, 13, 423-427. Ohashi, Y., Blamed. Mass Spectrom. 1984, 1 1 , 383-385. Ayanoglu, E.; Wegmann. A.; Pllet, 0.;Marbury, G. D.; Hass, J. R.; Djerassl, C. J . Am. Chem. Soc. 1984. 106. 5247-5251. Fenwick, 0. R.; Eagles, J.; Self, R. Biomed. Mass Spectrom. 1989, 10, 382-388. Jensen, N. J.; T o m , K. B.; Gross, M. L. Lip& 1066, 2 1 , 580-588. Jensen, N. J.; Tomer, K. B.; Gross, M. L. Lip& 1987, 2 2 , 480-489. Ross, M. M.; Nelhof, R.; Campana, J. E. Anal. Chlm. Acta 1986, 181, 149-156. Munster, H.; Budziklewlcz, H. RapM Gbmmun. Mess Spectrom. 1987, 1 , 128-128. Platt, J. A.; Uy, 0. M.; Heller, D. N.; Cotter. R. J.; Fenselau, C. Anal. Chem. 1988, 60. 1415-1419. Sweetman. B. J.; Tamura, M.; Hlgashlmorl, K.; Inagami, T.; Blak, I. A. Proc. ASMS Conf. Mass Spectrom. Allled Topics, 35th 1987, 508-509. Busch, K. L.; Cooks, R. 0. In Tandem Mass Spectrometry; McLafferty, F. W., Ed.; Wlley: New York. 1983; pp 11-39. Plattner, R. D.; Stack,R. J. Roc. ASMS Conf. Mass Spectrom. Allled Topics, 35th 1987, 520-521. Anhatt, J. P.; Fenselau, C. Anal. Chem. 1975, 47, 219-225. Huff, S. M.; Matsen. J. M.; Wlndlg, E.; Meuzelaar, H. L. C. B&med. Envkon. Mess Spectrom. 1986, 13, 277-286. Wlndlg. W.; Haverkamp, J.; Klstemaker. P. G. Anal. Chem. 1989, 5 5 , 81-88.
RECEIVED for review May 2, 1988. Accepted September 28, 1988. This work was supported by the US. Army CRDEC through US.Navy Contract N00024-85-C-5301and by the APL Independent Research and Development Fund. Mass spectra were obtained at the Middle Atlantic Mass Spectrometry Laboratory, a Shared Instrumentation Facility supported by Grants DMB 85-15390 and DMB 86-10589 from the National Science Foundation.
Flow Injection Gas Diffusion Method for Preconcentration and Determination of Trace Sulfide Emil B. Milosavljevie and Ljiljana Solujid Institute of Chemistry, Faculty of Sciences, University of Belgrade, P.O.Box 550, 11001 Belgrade, Yugoslavia James L. Hendrix* and John H. Nelson Departments of Chemistry and Chemical and Metallurgical Engineering, Mackay School of Mines, University of Nevada, Reno, Nevada 89557 A preconcentrating gas dtffusion flow injectlon analysis (FIA) method with amperometrlc detection has been developed for selective determination of sulfide. The method Is based on using the acceptor stream of the dlffuslon unit In a closedloop, moblie (recircuiating) mode for the accumulation of the analyte. I n the analysis step the contents of the acceptor loop are injected into a sodium carbonate stream and are carried to the amperometric detector. The various flow patterns requlred for this method are regulated by a single elght-port rotary valve. The method developed has very low detection ilmits of 0.15 pg/L. The relathre standard deviation for a 1 Hg/L standard Is 4 % , and of the potential interferences tested, only cyanlde had an effect.
Highly reproducible dispersion in flow injection analysis
(FIA)methods makes for convenient incorporation of sepa0003-2700/88/0360-2791$01.50/0
ration and/or preconcentration steps into the analytical cycle. Thus, both selectivity and sensitivity of the FIA methods are often substantially improved, compared to those of the classical procedures. Some of the possibilities of selectivity enhancement by FIA have been recently summarized (1). The two most studied methods for increasing the sensitivity of a given FIA method by preconcentration, with partial or total separation of the analyte from the matrix, are liquid/ liquid extraction and on-line use of ion-exchange columns. A recent review by Valcarcel and Luque de Castro (2),which contains 122 references, shows how both of these techniques have been successfully employed for solving numerous analytical problems. A preconcentrating step has been proposed for dialysis FIA (3). The method has been primarily developed for enhancing the selectivity of the anion-responsive electrodes in flow systems. It utilizes the upper channel of the dialyzer unit as an injection loop in a flow injection mode. The method was 0 1988 American Chemical Soclety
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ANALYTICAL CHEMISTRY, VOL. 60, NO. 24, DECEMBER 15, 1988
successful in improving the selectivity of the relatively nonselective anion-responsive liquid-membrane electrode. It also slightly improved the sensitivity at higher concentrations analyzed. However, probably due to the static nature of the acceptor stream, little or no improvement in the limits of detection has been realized. The probable reason for this phenomenon will be elaborated in the discussion section. A similar manifold has been developed for preconcentration and determination of cyanide by the gas diffusion colorimetric FIA (4).
A survey of the FIA methods for sulfide determination (5-16) shows that the ones with the best limits of detection utilize chemiluminescence (0.4pg/L (5) and amperometric detection (0.5 pg/L) (8). Both of these methods have good
precision and throughputa. However, the direct FIA/amperometry method for sulfide, without a preseparation step, would undoubtably suffer from numerous interferences. In this work a novel preconcentrating technique, based on the recirculating recipient (trapping) solution, was tested for the amperometric detection of sulfide. The aim was also to develop an FIA analytical system that would be less sensitive to interferences and have lower limits of detection than the existing ones. The concept presented and developed in this work can be easily extended for enhancing the sensitivity of the existing extraction, dialysis, other gas diffusion, and even liquid chromatography FIA methods.
EXPERIMENTAL SECTION The system manifolds, which are illustrated in Figures 1 and 3, were constructed from a FIAstar 5020 analyzer (Tecator, Inc.), equipped with two peristaltic pumps and a Tecator Chemifold V gas diffusion cell. The membrane used was a PTFE one supplied with the Chemifold, and it was changed every third day of operation. When the new membrane was installed, about 10 injections at 200 pg/L levels were needed to obtain reproducible signals (peak currents increased with the number of injections). The probable reason for this phenomenon is slight adsorption of hydrogen sulfide by a Teflon membrane. However, if adsorption does occur, it is irreversible under the experimental conditions used, and it did not interfere with sulfide determination even at submicrogram-per-literlevels. A variable volume injector L-100-1 (Tecator,Inc.) was used for a conventional gas diffusion manifold. A homemade eight-port valve similar to that described earlier (17) was used to control flow patterns in the preconcentrating gas diffusion manifold. All connections were made with either 0.51-mm-i.d. Microline or Tefzel tubing (1.02-mm id.). The accumulation loop for the preconcentrating system was made of Tygon pump tubing (0.38-mm i.d.), with connections to and from the valve and diffusion cell made of Tefzel tubing. These connections were made as short as possible to minimize the volume of the trapping solution (total volume of the accumulation loop, including the acceptor groove of the gas diffusion unit, was 130 pL);the sample loops were made from the appropriate lengthsof Tygon (1.02-mm id.) tubing. The flow-through amperometric cell (Dionex Corp.) described earlier (18) consisted of silver working and platinum counter electrodes. The reference electrode was a Ag/AgCl (1M NaCl), and it was separated from the flowing stream by an ion-exchange membrane. The silver working electrode was polished daily by using a small amount of toothpaste and a paper tissue. The flow-throughcell was potentiostated, and currents were measured by an Ion-Chrom/Amperometric detector (Dionex Corp.); the resulting signals were recorded on a strip chart Honeywell Electronic 195 recorder. Analytical reagent grade chemicals and deionized water additionally distilled from acidic potassium permanganate were used to prepare all solutions. A stock sulfide solution was prepared by dissolving 0.7490 g of a reagent grade Na2S-9H20(Fisher Scientific Co.) in distilledwater and then diluting this to 1L (100 pg/mL as S). The concentrationof thisstock solution was checked by the iodometric method. The standard solutions were prepared
. PO
Figure 1. Conventional gas diffusion FIA manlfold used for the determination of sulffde: (P) peristattic pump; @) dmusion -#; (I) lnvalve; (MC) mixing coil; (FC) flow-through cell; (PO) potentlostat; (w)
waste. Flow rates are given in milliliters per minute. See text for details. A
A
900-
- 800.2
;r
700
-
600.a 500'U
2 400-
P
3001 0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
INJECTED VOLUME (mL 1
t /
L 1
2
3
4
5
INJECTED VOLUME (mL1
Figure 2. (A) Variation of peak currents with InJected volume for the conventional gas diffusion FIA manlfold (200 pg/L sulfide standards were injected). (B) Variation of peak currents with Injected volume for the preconcentratlng gas diffusion FIA manifold (20 pg/L sulfide standards were Injected).
daily, except for low-concentration work, when the standards were diluted prior to each analysis. In order to minimize hydrogen sulfide evolution, all sulfide standards were made to be 1 X lo4 M in potassium hydroxide.
RESULTS AND DISCUSSION Performance and Characteristics of the Conventional FIA Gas Diffusion Manifold. In order to compare the preconcentrating method with a conventional gas diffusion FIA, the latter one was first optimized for the determination of sulfide. The manifold used, which is similar to the ones described previously (19-21), is shown in Figure 1. The effects of several parameters on the performance of this analytical system were studied. Experiments were carried out to establish the effects of the injected volume, with the results shown in Figure 2A. As was seen before (22),sample volumes greater than 0.5 mL caused the system to approach a steady state, without appreciably increasing the sensitivity of the system. The effect of the applied potential at the working silver electrode was investigated in the range -0.20 to +0.40 V vs a Ag/AgCl reference electrode. The optimum potential found
ANALYTICAL CHEMISTRY, VOL. 60, NO. 24, DECEMBER 15, 1988
was 0.00 V. Slightly lower responses were obtained at potentials in the range f0.20 V. However, at potentials above 0.30 V the background noise was too high. When one is selecting the mixing coil dimensions, there is an obvious trade-off between better acidification and dilution of the sample. The former enhances the analytical signal, and the latter attenuates it. The optimum mixing coil dimensions were found to be 30 cm X 0.7 mm i.d., and in all subsequent experiments that size coil and a potential of 0.00 V were used. The temperature chosen for this study was 30 "C.The current response at 30 "C was approximately 20% greater than at 20 O C . At temperatures above 40 O C bubble formaton substantially increased the background noise. The four acceptor solutions tested were 0.01 M sodium carbonate, sodium hydroxide, and potassium nitrate, as well as distilled water. As expected, the first two gave the best results, and sodium carbonate was chosen for further work. Various concentrations of sulfuric acid used for carrier (C) and reagent (R,)were tested. For concentrations greater than 0.1 M HaO,there was no change of the response current when a given sulfide standard was injected. Also, higher currents were recorded when both the acceptor and donor streams were flowing in the same direction compared to counter stream flow. Particularly interesting are the results of the experiments when the effect of the flow rate of the acceptor stream was investigated. It is generally accepted that the amperometric current (I)measured in a flowing stream is related to the flow rate (V) and concentration (C) by the following equation (see for example ref 23):
I = nFAkVaC where a is a constant that depends on the electrode geometry and k is a constant related to the diffusion coefficient and geometric parameters; n, F,and A have their usual electrochemical significance. Therefore, the peak height (current) should increase with increase in flow rate. Our data are just the opposite. The current response to an injection of the s a l d e standard, throughout the interval investigated (0.15-1.2 mL/min), decreased in a nonlinear manner with an increase in the flow rate of the acceptor stream. In the only other study of the FIA gas diffusion system with amperometric detection (241, it was pointed out that the above equation does not take into consideration phenomena such as the adsorption mechanisms and redox kinetics at the surface of the electrode. These effects could be particularly important when the working electrode is itself oxidized, as is the case with sulfide determination when a silver electrode is used. Also, when the ratio of the donor and acceptor stream flow rates is greater than one, a preconcentration of the analyte in the acceptor stream occurs. The preconcentration factor increases with M increase in this ratio. In other words, for a cbnstant donor stream flow rate, the amperometric signal would increase with a decrease in the flow rate of the acceptor stream. For subsequent experiments a compromise flow rate of 0.3 mL/min for the acceptor stream was chosen since lower flow rates would make the analysis time too long. The slope and correlation coefficient for the 0.02-1 pg/mL calibration interval were 1.38f 0.04pA/&g/mL) and 0.9995, respectively. The corresponding data for the 0.5-5 pg/mL range were 0.91 f 0.34 pA/(pg/mL) and 0.9909. It may be seen that the sensitivity decreases with an increase in concentration and that at higher concentrations the linearity is preserved in the concentration interval spanning just 1order of magnitude. In the only other FIA gas diffusion study with amperometric detection (24),the Calibration graphs had two linear ranges, 0.06-6 and 12-110 pg/mL, the slope for the higher concentration range being about 2 orders of magnitude lower. We do not know the real reason for such behavior of the amperometric detector. It could be that at higher con-
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VALVE POSITION
H,SO,(RJ Na,CO,(R,)
H,SO,(C)
MC VALVE POSITION
P
B
H X 4 (R,)
S
L MC
a
-
Figure 3. Schematics of the preconcentrating gas diffusion F I A manifold used for the determination of sulfide: (P) peristaltic pumps: (D) diffwlon cell;(MC) mixing coil; (FC)Row-through cell; (PO)potenthtat; (S) sample: (L) sample imp: (AL) accumulation bop: (6) b W e d ports: (w) waste. The description of the valve functionsand other details are
given in the text.
centrations (in our system above 1pg/mL) there is a change in the electrodissolution kinetics of the working silver electrode, which causes a decrease in the electrolysis efficiency. The other FIA gas diffusion methods utilizing other types of detectors have shown dynamic linear responses in the concentration intervals spanning 3 or 4 orders of magnitude. From the aforementioned, it may be safely concluded that the unusual linear ranges are detector-induced. With the conventional FIA gas diffusion manifold described in this section the relative standard deviation (n = 10) for a 0.5 pg/mL standard was 1.3%. The limit of detection for this analytical system was 4 pg/L, and 40 samples could be analyzed in 1 h. Performance and Characteristics of t h e Preconcentrating FIA Gas Diffusion Method. The manifold developed (Figure 3) is based on using the acceptor channel of the diffusion unit in a closed-loop, recirculating mode for the accumulation and preconcentration of the analyte. In this manner, part of the analyte present in the large sample volume (for example L = 5 mL) is preconcentrated into the much smaller volume of the trapping solution (in all the experiments with this manifold the accumulation loop had a volume of 130 PL). As may be seen from Figure 3,the various flow patterns required for this analytical system are regulated by a single eight-port rotary valve. When the valve is in position A (accumulation/preconcentration step, Figure 3A), the sample plug in the sample loop L is washed by the sulfuric acid carrier (C) to a mixing point with the reagent (Rl), which is also 0.1 M sulfuric acid. The mixing coil (30 cm X 0.7mm i.d.), which ensures thorough acidification of the sample, is positioned
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ANALYTICAL CHEMISTRY, VOL. 60, NO. 24, DECEMBER 15, 1988
downstream. The acidified sample then passes through the donor channel of the diffusion unit and goes to waste. At the same time the 0.01 M sodium carbonate trapping solution is being recirculated through the acceptor channel of the diffusion unit. Hydrogen sulfide formed in the donor stream diffuses through the Teflon membrane and is converted to the sodium salt in the acceptor channel. The preconcentration occurs since the volume of the sample loop (L) is larger than the volume of the accumulation loop (AL). Sodium carbonate (R,) flows through the amperometric detector at a flow rate of 0.3 mL/min. The sample flows to waste, or in order to lower sample consumption, the sample pump may be stopped when the valve is in position A. When the valve is switched to position B (analysis step, Figure 3B), sodium carbonate (h) washes the contents of the accumulation loop (AL) to the flow-through amperometric cell (FC)where sulfide is detected. Both the accumulation loop and R2are positioned on the same pump head and have the same flow rates (0.3 mL/min), which ensures a constant flow through the flow cell. This is important when the amperometric detection is used, since changes in the flow rate cause changes in the base line. Looking at Figure 3, one might conclude that the flow rates through the detector in valve positions A and B are 0.3 and 0.6 mL/min, respectively. However, this is not the case. The pump tubing of the accumulation loop, along with the pump rollers, does not allow an increase in the flow rate when the valve is switched from the position A to B. Thus, irrespective of the valve position, the flow rate through the amperometric detector is constant. Also, when the valve is in the analysis position, the sample pump is switched on and the new sample fills the loop L for the subsequent analysis. Experiments were carried out with different sample loop volumes. The results obtained when various sample volumes of a 20 pg/L sulfide standard were injected into the donor stream are shown in Figure 2B. As may be seen, there is a linear relationship between the sample volume and the peak current. The intercept, slope, and correlation coefficient for this line were -0.01 f 0.04 pA, 0.112 f 0.041 pA/mL, and 0.9985, respectively. One may see that no steady-state signal is achieved even for the large sample volumes, and that the sensitivity of the analytical system is directly proportional to the sample loop volume. This, of course, was not the case with the conventional gas diffusion manifold (see figure 2A). It should also be noted here that a good linear correlation between the sample volume and the response of the detector enables the calibration runs to be performed with any sample volume, as long as the analyte concentrations do not approach the limits of detection for a given sample volume. An additional advantage of the manifold with the recirculating acceptor solution compared to the conventional gas diffusion one is the extended linear range. For example, when a 1-mL sample loop was used, the linear response of the amperometric detector spanned 3 orders of magnitude (1pg/L to 1 rg/mL). The slope and correlation coefficient for this calibration run were 4.3 f 0.3 pA/(pg/mL) and 0.9981, respectively. As expected, higher sensitivities are obtained for greater sample volumes. The slope and correlation coefficient for a calibration run with a 5-mL sample loop were 18.5 f 0.5 nA/(pg/L) and 0.9990, respectively. However, at submicrogram-per-liter levels the sensitivity decreases. For the range (0.25-1 pg/L) the relevant data are 14 f 1 nA/(pg/L) and 0.9989. At these very low concentrations the solubility of hydrogen sulfide in 0.1 M sulfuric acid (donor stream) probably begins to play a significant role. In the preconcentrating system the ratio of the sample loop and accumulation loop volumes is about 38, when L = 5 mL.
IOmin c-----*
A
J
U
40 I
B
SCAN
+
Figure 4. (A) Recording of five successive injections of a 1 pg/L
standard (5-mL sample loop). (e) RapM-scan response to an InJeCtion of the same standard (valve switchings are indicated by the arrows).
However, this is not a preconcentration factor, since not all of the hydrogen sulfide formed in the donor stream diffuses through the membrane. Therefore, calculating the preconcentration fador is not as straightforward as with FIA systems that contain ion-exchange columns for preconcentration of the analyte. The gas diffusion systems always sacrifice sensitivity to achieve better selectivity. Perhaps the best comparative parameter between the conventional and preconcentrating FIA gas diffusion methods can be obtained by comparing the slopes of the calibration graphs and correcting for the sample volume that is seen by the detector (in our system 100 and 130 pL, respectively). Such a parameter, which might be called “the apparent preconcentration factor”, has a value of 17 for our system. The relative standard deviation (n= 5) at 1pg/L levels was 4%. A tracing of five successive injections of this standard is shown in Figure 4A. The spikes in the tracing are caused by the slight flow disturbances caused by valve switching. As is better seen on the rapid-scan tracing (Figure 4B), these spikes are removed from the FIA signals and do not interfere. From this figure it may also be seen that the peak heights slightly but steadily decrease (the rapid scan was recorded after five successive injections of a 1pg/L standard). This is probably due to the oxidation of sulfide, even though all precautions were taken to deaerate the water used for making the standards. The limit of detection for the preconcentrating analytical system with the 5-mL sample loop, calculated according to a recent recommendation (W), is 0.15 pg/L. Thus, the method has, as far as we know, lower limits of detection than any of the existing FIA methods for this analyte. From the description of the manifold it is obvious that the higher sensitivity is achieved at the expense of the sample throughput. For example, with 1-mL and 5-mL sample volumes, 20 and nine analyses per hour could be performed, respectively. It is also obvious that even higher sensitivities may be achieved by increasing the sample loop volume. In this study we have also tried to compare the manifold described in the present work with one that utilizes a static acceptor solution suggested earlier ( 3 , 4 ) . However, unusual signals were obtained with the latter. Rapid-scan tracings of the amperometric detector response when 1 and 5 pg/L standards were injected are shown in Figure 5. The double peaks obtained point to the lack of analyte in the center of the sample zone (26). The probable explanation for this
ANALYTICAL CHEMISTRY, VOL. 60, NO. 24, DECEMBER 15, 1988
2705
Table I. Solutions Tested for Their Possible Interference" compd
concn, M
compd
NaCl NHJ KSCN NaNOP MgC12 Na2S04
1.0 0.1 0.01 0.001 0.1 0.1
KBr NazSzOs NazCOs NazSOs NaCH,COO Na2EDTA
concn, M 0.1
0.001 0.01 0.001 0.1 0.1
@Theresponse of the amperometric detector to injections (5 mL) of the solutions tested could not be distinguished from the background noise.
I
SCAN
--*
Figure 5. Rapid-scan response curves obtained by injection of (A) 1 pg/L and (6)5 pg/L sulfide standards using a manifold with the static
acceptor solution (valve switchlngs are indicated by the arrows). phenomenon lies in the difference of the two approaches. In the manifold with the recirculating acceptor solution there is an even distribution of the diffused analyte throughout the accumulation loop. On the other hand, when a static acceptor solution is used, only the portion of this solution that is in direct contact with the dialysis or diffusion membrane traps the analyte species by the pH, redox potential, or some other change relative to the donor stream. Transfer of the analyte a c r w the membrane and ita trapping in the acceptor solution, by one of the possible mechanisms, are equilibrium processes. Thus, the shape of the signal obtained could very well be explained by the partial back-diffusion of the analyte from the acceptor solution to the donor stream. In other words, the distortion of the signals, when the static acceptor solution is used, is caused by the incompleteness of the trapping process. This might be accentuated since during the preconcentration step the concentration of the analyte in the acceptor solution increases. There is another plausible explanation for the behavior of the system with the static acceptor solution. The analyte transfer across the membrane is governed by the following processes (21): diffusion of the analyte from the bulk to the solution/membrane interface, partition between donor solution and membrane phase, diffusion inside the membrane, partition between the membrane and acceptor solution, and diffusion from the surface into the bulk of the acceptor phase. The recirculating flow of the acceptor solution surely influences the last, and maybe even the last two, processes. On the other hand, when the stationary acceptor solution is used, it is possible that some of the hydrogen sulfide is held in the membrane pores, or at the interface, and that there is not sufficient agitation to complete the trapping process. One should not forget that we are dealing with very low concentrations of the anal@. There are several possible reasons why other authors ( 3 , 4 )have not noticed this phenomenon, two of which are higher analyte concentrations (limits of detection) and different detection methods. Also, it might be that these processes are analyte-related. However, in relation to the applicability of the method, when a recirculating acceptor solution is used, all the processes causing distortion of the signal shape are either completely eliminated or are substantially attenuated. Table I summarizes the study of possible interferences. This study was carried out on a preconcentrating system with a 5-mL sample loop volume. It has been established previously that the PTFE membranes used in the FIA gas diffusion studies are effective barriers for ionic species (21,221. Nevertheless, some of the anions, which would interfere in the
determination of sulfide with the amperometric detector if no membrane were used, were tested anyway (Cl-, Br-, I-, SCN-, Sz032-). The concentrations of these species given in Table I are the maximum concentrations at which they were tested. As expected, in all these cases the response of the amperometric detector could not be distinguished from the base line. The other major group of anions ( N O , SO3*, COSs, S2032-)were tested since, when acidified, they form acidic gaseous oxides that do diffuse through the membrane. These acidic oxides at sufficiently large concentrations, even though they are not electroactive a t the working potential, could interfere in a roundabout manner. The Ag/AgCl reference electrode in the configuration used is separated from the flowing stream by the ion-exchange Ndion membrane. If the buffer capacity of the acceptor solution is too low, there is a significant change in pH when the acidic oxides diffusing through the membrane are trapped in this solution and the potential of the reference electrode assembly changes. This change is probably induced by a shift in the ion-exchange equilibria at the Nafion membrane. The concentrations of the anions forming acidic gaseous oxides given in Table I are the maximum concentrations that could be present in the sample and not be distinguished from the background noise, when 0.01 M NaZCO3was the recirculating acceptor solution. The interferences from even higher concentrations of COZand NO2- (0.1 M) correspond to only 0.75 and 3 pg/L sulfide (apparent). Thus, a very good selectivity factor of about 1 x lo6is achieved. An even better selectivity factor is achieved when a more concentrated acceptor solution with a higher buffer capacity is used (0.1 M KOH). The only serious interferant found for this system was cyanide ion, which is preconcentrated and determined in a manner analogous to that for sulfide. However, the preconcentrating manifold developed is compatible with any detection mode used for FIA, and when both sulfide and cyanide are present in the same sample, a spectrophotometric detector with a selective chromogenic reagent for sulfide could be used. We plan another approach the development of a manifold, which would incorporate an anion-exchange column just upstream from the flow-through cell, for the preconcentration, separation, and subsequent determination of both of these species. Comparison of the sulfide concentrations determined by the preconcentrating and conventional FIA gas diffusion methods gives the following linear regression parameters: slope, 1.001 f 0.003; correlation coefficient, 0.9984. The corresponding values for the comparison with the ion chromatography method (18) are 1.02 f 0.07 and 0.9980.
CONCLUSION Higher sensitivity and lower limits of detection are the most important advantages of the preconcentrating FIA gas diffusion method over the conventional one. These advantages are a consequence of the fact that no steady-state signals are obtained even for the very large sample volumes, so that the sensitivity of the preconcentrating method is directly pro-
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Anal. Chem. lB88, 60,2796-2801
portional to the sample volume. The analyte is effectively preconcentrated into the much smaller volume of the recirculating acceptor solution in which the analyte is subsequently determined by the amperometric detection. Since a single eight-port, two-position valve is used to control all the flow patterns in the preconcentrating gas diffusion manifold, the system may easily be fully automated and miniaturized. As a consequence of the preconcentrating/accumulation step, the preconcentrating method has a lower throughput than the conventional one. Registry No. Sulfide, 18496-25-8.
LITERATURE CITED Pacey, G. E.; Hollowell, D. A.; Miller, K. G.; Straka. M. R.; Gordon, G. Anal. Chlm. ACte 1988, 179, 259. Valdrcei, M.; Luque de Castro, M. D. J . chrometogr. 1987. 393, 3. Martin, G. B.; MeyemOff, M. E. Anal. C h h . Acta 1988, 186, 71. Zhu, 2.; Fang, 2. Kaxue Tm@o (Forelgn Le-. M.)1888, 31, 1728. Burguera, J. L.; Townshend, A.; (Leenfleld, S. Anal. Chlm. Acta 1980, 174, 209. Duffleid, E. J.; Moody, G. J.; Thomas, J. D. R. Anal. plot. 1980, 17, 533.
L&Wtt, D. J.; Chen. N. H.; Mahadevappa, D. S. Anal. Chim. Acta 1981. 128. 163. Me, H.; Yan, H. Kexue Tongb80 (Forelgn leng. Ed.) 1982, 2 7 , 959. Bablker, M. 0.;Delzlel, J. A. W. AM/. Roc. 1983, 20, 609. Bwguera, J. L.; Burguera, M. Anal. Chh.Acta 1884, 157. 177. Rbs, A.; Luque de Cestro, M. D.; Vaicircel, M. Aneiysf (bndon) 1984, 109, 1487.
(12) Glaistef, M. G.; Moody, G. J.; Thomas, J. D. R. Analysf (London)1985, 110, 113. (13) Kurzawa, J. Anal. Chlm. Aeta 1885, 173, 343. (14) Johnson, K. S.; Beehler, C. L.; Sakamoto-Arnold, C. M. Anal. Chhn. Acta 1988, 179, 245. (15) Peterason. B. A.; Fang, 2.; RuFlEka, J.; Hansen, E. H. AMI. Chim. Acta 1888, 184, 165. (16) Dasgupta, P. K.; Yang. H. C. Anal. Chem. 1988, 56. 2839. (17) Fang, 2.; RElEka, J.; Hansen. E. H. Anal. Chlm. Acta 1984, 164, 23. (18) Rocklin, R. D.; Johnson. D. C. Anel. Chem. 1983, 55. 4. (19) BaadenhulJsen,H.; SewenJacobs, H. E. H. C h . Chem. (W/nsfon-Salem, N . C . ) 1979, 25, 443. (20) RiZiEka, J.; Hansen, E. H. Flow Injecfbn Analysk; Wliey: New York, 1981. (21) Van der Linden, W. E. Anal. CMm. Acta 1983, 151, 359. (22) Hollowell, D. A.; Pacey, G. E.; Gordon, 0. Anal. Chem. 1985, 57, 2851. (23) MacKoul, D.; Johnson, D. C.; Schick, K. G. Anal. Chem. 1984, 56, 436. (24) Granados, M.; Maspoch, S.; Blanco, M. Anal. chim. Acta 1988, 779, 445. (25) Analytical Methods Commltiee. Analyst (Lonrkn)1887. 712, 199. (26) RQiiEka, J.; Hansen, E. H. Anal. Chlm. Acta 1986, 780, 41.
RECEIVED for review September 14,1987. Resubmitted June 27,1988. Accepted September 15,1988. This research was supported by the United States Bureau of Mines under the Mining and Minerals Resources Research Institute Generic Center Program (Grant Number G1125132-3205, Mineral Industry Waste Treatment and Recovery Generic Center).
NanoHter Volume Sequential Differential Concentration Gradient Detector Janusz Pawliszyn Department of Chemistry, University of Waterloo, Waterloo, Ontario, Canada N2L 3Gl
A sknple low volume dlfterentlal refractlve index gradient detectlon scheme has been described. Thls method elbnlnates low-frequency ndse associated with refractive Index drMs produced by temperature hmtabllltles andlor gradient solvent progremmlng condnkne. The reduction of laser posItion nolse and mechanical vlbratlons was also achleved by anoveloptkalmangemecrt. lBedetectorcomlstsdadngle He-Ne laser, a slmple 0pUcal procebdng scheme,and a single photosensor. The sdgnal produced by thls 881uor Is proportknal to concentratlon by over 4 orders of magnitude with the rdractlve Index gradlent detectlon h i t of 2 X lo-' r e fractlve Index unltlm. TMs translated into a concentration ddectbn Unn of 7 X lo-' M sucrose (subpicogram level) for our detector design. The detectkn volume Is about 2 nL and can be made even smaller by focusing a laser beams dhectly Into the caplllary. This scheme can be used both as a unlversai method (slgnai proportional to the second derivatlve of the concentratlon In respect to distance) or as an optlcal absorption probe (through photothermal process, In which case the signal Is proporttonal to the second derivative of optical denrlty In respect to position). Thls sensor has been appbd In flow lnjectlon analysis and caplllary zone electrophoresls.
The introduction of capillaries to separation technology has resulted in the generation of high efficiency methods (I). They 0003-2700/86/0360-2796$01.50/0
are characterized by smaller sample volumes, higher resolution, and shorter separation times compared to conventional techniques. Capillary chromatography is already well accepted when gases (2) or supercritical fluids (3) are used as mobile phases. Recently, much research effort has been directed to designing systems involving liquids such as in capillary liquid chromatography (4) and capillary zone electrophoresis (5,6). These schemes require very small inside diameter capillaries to operate at optimum efficiencies, sometimes smaller than 10 Mm i.d. (7).This development necessitates the design of ultralow-volume detection methods that are able to monitor narrow bands within small bore capillaries. Many techniques developed for this purpose have been employing lasers as light sources (8). In most cases, well-accepted detection schemes have been used. Laser beams, with their collimated characteristics, allow fine focusing of high-intensity beams and therefore reduction of detection volume by several orders of magnitude. Capillary separation methods using the liquid mobile phase are able to achieve close to a million theoretical plates per meter of capillary (9). They are likely to separate even quite complex mixtures. Therefore, the ideal detector for capillary methods would be a nondestructive universal type that is able to monitor all eluting components. If full separation cannot be achieved or additional information about the sample is required, then simultaneous selective analysis can follow universal detection. One detector type, which fulfills these requirements, applies refractive index monitoring. Much effort has been recently placed on developing an ultrasmall 0 1958 American Chemical Society
