Anal. Chem. 1995, 67,2110-21 18
TwomDimensionai Conductometric Detection in ion Chromatography. Analyte identification, Quantitation of Very Weak Acid Anions, and Universal Calibration Anna SJOgren and Pumendu K. Dasgupta* Department of Chemistry and Biochemistry, Texas Tech Univetsity, Lubbock, Texas 79401-1061
A universal and sensitive detection method for anion chromatography is described. Following suppressed conductometric detection canied out with an NaOH eluent and an electrical suppressor, the eluite is transported through a microscale electrodialyticNaOH generator and a second conductivity detector. While the first detector responds primady to strong acid anions, the second detector records a decrease in the NaOH background signal, regardless of the p& of the analyte. Routine detection of very weak acids at the low micromolar level is possible. The dual detection method not only constitutes the most generally applicable approach to IC by combining the detection merits of both suppressed and single-columnIC, it provides possibilities of peak identiiication beyond retention characteristics. Area ratio values from the two detector signals provide unique signatures for each analyte ion. It is possible to estimate the limiting equivalentconductance of an unknown eluite and the analyte concentration without specific calibration. The method is also effectivein diagnosingthe occurrence of coelution. A situation common to all chromatographicmethods utilizing bulk property detectors is that the retention time of an eluite constitutes merely an indication of its identity. Along with refractometry, conductometry is among the most general of all liquid-phase detection methods. Conductometry is ideally suitable for ion chromatography (IC): electrical conductivity is a property shared by all ions and uncharged species cannot interfere. In 1975, Small et ale1developed a method that eliminated the eluent conductivity in a postseparation H+exchange step termed suppression; the decrease in the background conductance allowed dramatically improved strong acid analyte detection limits. Subsequent refinements to the technique, such as electrodialytic suppression and eluent improved column technolo& and system hardware, have made possible present day systems that can easily detect analyte signals corresponding to a (1)Small, H.;Stevens, T. S.; Bauman, W. C. Anal. Chem. 1975, 47, 18011809. (2)Strong, D. L.; Joung, C. U.; Dasgupta. P. K j . Chromatogr. 1991,546,159173. (3) Strong, D. L.; Dasgupta, P. K Anal. Chem. 1989,61,939-945. (4)Strong, D. L.;Dasgupta, P. K; Friedman, K; Stillian,J. RAnol. Chem. 1991, 63,480-486. (5) Rabin, S.; Stillian, J.; Barreto, V.; Friedman, K; Toofan. M. j. Chromatog. 1993,640,97-109. (6) Stillian, J. R; Pohl, C. A j. Chromatogr. 1990,499,249-266.
2110 Analytical Chemistry, Vol. 67, No. 73,July 7, 7995
change in conductance less than a tenth of the conductance of pure water. Suppressed conductometric IC has been a resounding success in both research laboratories and the marketplace. However, for weak and especially very weak acid anions, poor sensitivity and markedly nonlinear calibration behavior constitute significant shortcomings. Also, while the technique is generally applicable for strong acid anions, it is a problem to identify the eluted ions beyond their retention behavior. Ancillary confirmation of the identity is always desirable: the validation of analytical data is increasingly important. Efforts to achieve high and relatively uniform sensitivities for all analytes have included various manipulations of the suppressed eluate.7S8 Identification efforts have included the sequential conductance measurement of the nonsuppressed and the column-suppressed e l ~ a t e .On-line ~ conversions of the analyte, first from an acid to a base and then from an acid to a salt,have been described in a series of papers.'O-12 These e f f ~ r t s are ~ - ~all~ based on dual conductometric detection. New clues to the eluite identity are obtained with two signals that can be analyzed both independently and in combination. Serial indirect conductometric and UV detection have also been used to deduce the identity of an eluite Compared to conventional suppressed IC, another major advantage of these systems is that weak and very weak acids can also be detected. This paper presents a complete theoretical background and experimental refinements of the most successful of these approaches: NaOH eluent suppressed conductometric IC followed by on-line electrodialytic NaOH introduction and conductometric detection12to provide universal calibration, estimation of equivalent conductance/ pKa values, diagnostics for peak coelution, and compatibility with gradient elution. OPERATINO PRINCIPLES When NaOH is used as the eluent, after suppression the background should theoretically be pure water. In practice, the suppressor background effluent is slightly acidic because of the presence of anionic impurities in the eluent and the intrusion of (7)Downey, S. W.;Hieftje, G. M. Anal. Chin. Acta. 1983, 153,1-13. (8)Shintani, H.; Dasgupta, P. K Anal. Chem. 1987,59,1963-1969. (9)Sato, H.; Shishido, Y.; Furuya, K; Dasgupta, P. IC, unpublished studies, 1992. (10)Berglund, I.; Dasgupta, P. K. Anal. Chem. 1991,63,2175-2183. (11) Berglund, I.; Dasgupta, P. K Anal. Chem. 1992,64,3007-3012. (12)Berglund, I.; Dasgupta, P. K; Lopez, J. L.; Nara, 0.Anal. Chem. 1993,65, 1192- 1198. (13)Sato, H.Keynote lecture presented at the International Symposium on Ion Chromatography, Turin, Italy, September 1594. 0 1995 American Chemical Society 0003-2700/95/0367-2110$9.00/0
omnipresent COZ. Since un-ionized eluites are invisible to a conductivity detector, the undissociated fraction of any weak acid eluite will not elicit a detector response. The signal is thus strongly dependent on the pKa of the eluite. This essentially means that eluite acids with pKa 2 7 cannot be detected by suppressed IC at all (PKa 2 6 for a carbonatecontaining eluent) and the linear dynamic range for the determination of eluite acids in the pKa range 3-6 is quite limited. If, however, the detector influent background was alkaline instead of slightly acidic, all but eluites of the highest pKa values will be ionized and will thus be sensed by conductance detection. If the suppressor consistently suppressed exactly 99.9%of a 100 mM NaOH eluent, the detector influent will contain 0.1 mM NaOH (PH 10) and acid eluites with a pKa value of 5 -10 will be signifmntly ionized. While there is no known method to achieve an exact and constant degree of substoichiometric neutralization, it is possible to introduce a small and constant quantity of NaOH after quantitative neutralization and prior to the measurement of conductance. The concentration of NaOH should be limited to avoid a large (and noisy) detector background. As previously shown,lZthe introduction of a small constant amount of NaOH is best accomplished by electrodialysis. As an eluite HX comes through, the following reaction occurs: OH-
+ HX
4
X- + H 2 0
is C molar, is given by
Where dH and IX are the equivalent conductance of H+ and X-. For a weak monoprotic acid, the corresponding signal GI, is given by
where C is given by
+ 4KC91'2)
C = 0.5(-K+ (K2
(4)
where K is the dissociation constant of acid Hx The above expression for GI, is an approximation; it is not applicable at extreme dilutions or for values of K so low that the dissociation of HX is affected by the traces of acid present in the background suppressor effluent. The general equation for weak polyprotic acids can be formulated as
(1)
with the result that OH- is replaced by an equivalent amount of X- and because the equivalent conductance of OH- is greater than that of X-, a decreased conductance (i.e., negative) signal results. Obviously, this response is not particularly analyte specific (except that extremely weak acids, pKa 2 11,are still largely undetected) and is essentially governed by the same detection principles that would be operative for an NaOH eluent, unsuppressed IC system.14 The latter system, however, is largely impractical because the full concentration of the NaOH eluent appears as the detector background. In contrast, in the system discussed here, the detector NaOH background has no direct relationship with the eluent NaOH concentration and the user is free to choose any value for either the eluent or the detector background NaOH concentration. The experimental arrangement is largely the same as that described previously.lZ An NaOH eluent anion chromatography system is configured with a water regenerant electrical suppressor that is followed by the first conductivity detector (Dl). The background effluent from this detector is nearly pure water, containing a trace amount of acids resulting from non-hydroxide anions present in the eluent. This is followed by the microelectrodialytic NaOH generator (MENG) and a passive low-dispersion mixing device to achieve mixing of the electrodialytically introduced NaOH and the D1 effluent. A second conductivity detector 0 2 ) then monitors the conductance again. THEORETICAL CONSIDERATIONS
The following embodies generally applicable theoretical considerations for our experimental system. In specific cases, aprroximations and simplifications are possible; see, e.g., refs 1@ 12. For a strong monoprotic acid HX, the D1 signal Glsm(ii pS/ cm) at any point in the eluite band where the eluite concentration
where IH(n-i,X is the equivalent conductance value of the ion Hn-;Xi- and the a values can be expressed as n
ai = (KO, ...,K i [ H + ] n - i ) / ~ ( K o..., , Ki[H+ln-i)
(6)
I=O
KI through K,, are the first through nth dissociation constants of H,,X and KOhas a value of 1. A polynomial in [H+l, with a single unique real solution, is constituted by the charge balance expression n
[H'] = c i C a i
(7)
i= 1
With most polyprotic acids, the above expressions can be simplified in practice, some dissociations being negligible and other, closely spaced dissociation steps occurring to comparable degrees. The second detector background, (&bad, is dependent on the MENG current. It can be expressed as G2b-d
@S/Cm) = 1000(60Ei/96500&)
AN^ + &H)
(8)
where E is the faradaic efficiency (most MENG designs operate with near-unity faradaic efficiency), i is the MENG current (in mA), and Q is the chromatographic flow rate (ii mWmin). Substitution of numerical values and consolidation of these lead to a simpMed form of eq 8 that directly relates the D2 background to the MENG current, device efficiency, and the chromatographic flow rate:
~~
(14) Okada, T.; Kuwamoto, M. Anal. Chem. 1985, 57,829-833.
Analytical Chemistry, Vol. 67, No. 13, JuW 1, 1995
21 11
The exact background conductance will still depend on the extent of impurity anions present in the eluent. However, unless the amounts of impurities present are very significant, the precise value of Gbgnd does not control the magnitude of the eluite signal; its choice is only important to us to reach an optimum value between the conDicting demands of having sufticient free NaOH available to equal or exceed the maximum eluite concentration expected and keeping the concentration low enough to maintain GZbgnd low and thus minimize detector noise. A Gbgnd value of 20-35 pS/cm (130-225 pA MENG current for unity efficiency at an eluent flow rate of 1mL/min) is optimum for most purposes, providing a background NaOH concentration of -100 pM (accounting for typical chromatographic dilution, this will accommodate injected analyte concentrations of 1mN for each separated analyte). For monoprotic acids with pKa I8 and for n-protic acids with pKa, being 18, neutralization of the eluite acid by the NaOH introduced is complete as long as modest concentrations are injected. The signal G at any point in the eluite band, where the eluite concentration is C molar, is given by G2, = 1 o o o c ( A O H - AX)
(10)
For the simplest case of a strong monoprotic acid, if the two detector signals, (eq 2) and Gz, (eq 10) are plotted against each other, the slope S ,, (or the ratio of Glsm to G d is constant, independent of the eluite concentration, and given by
Considering that AX values for common anions range approximately from 30 to 80, S ,, will range from 2.25 to 3.63. A rearranged form of eq 12 is useful for determiniig AX:
- AH)/(ssm
AX = (s&OH
+ l)
(13)
The caveat in determining IX is to be cognizant of the uncertainties caused by the propagation of errors; for an intermediate mobility anion of AX = 50, an uncertainty of 2% in determining GI,, and Gz, translates to an uncertainty of 2.8% in Ss, and thence to -10% in determining AX. For the weak monoprotic acid case, the slope,S is going to be concentration dependent, decreasing with increasing C. Dividing eqn 3 by eq 10 yields , ,S
= (0.5SS,(-K
+ (K' + 4KC)'/'))/C
= 0.5Ss,K/C
The behavior is similar to that of strong monoprotic acids in that the slope is basically governed by the ionic mobilities. However, in this case not only the equivalent conductance of the monoanion but also that of the fully ionized polyanion is involved. Note that although Axn- is generally substantially higher than A H ( ~ - ~ ) xthe , value of Slspwill typically be lower than or, at best, equal to the range of values observed for S ,, in eq 12 because of the appearance of the factor n in the denominator of eq 17; i.e., the analyte exhibits a greater number of equivalents in D2 than in D1. Atypical case where eq 17 applies is that of orthophosphate as analyte, the third proton of H3P04not playing any discernible role. At typical conditions used in IC, it is fully dissociated to HzP04- in D1 with essentially no dissociation to H P O F while in D2 the only anionic form of the analyte is HPOP. In many other cases, there is significantly incomplete dissociation in D1 even for the first proton. In such a situation, it is necessary to modify eq 17:
and the exact value of ,S will exhibit a dependence on C similar to that previously discussed for eq 14. One final case merits an individual discussion, that of weak acid analytes with pKa 2 8. These analytes may be incompletely ionized with a background NaOH concentration of 0.1 mM. It is sufficient to consider the case of a monoprotic acid because subsequent dissociation steps with pKa values even higher are likely to contribute very little to the observed signal. For such analytes, e.g., borate, cyanide, silicate, etc., there is no discernible signal in D1. Consequently, the GI/& quotient is essentially zero. In the second detector, only the fraction that is ionized contributes to the D2 signal and this is a function of the amount of NaOH introduced through the MENG. The amount of NaOH introduced is available by appropriately modifying eq 9:
[Na'] = 0.623Ei/Q
(15)
With the approximation that, in comparison with OH-, H+ is negligible in the second detector (this is reasonable under all conditions in which the amount of eluite acid does not overwhelm the amount of NaOH introduced), the D2 signal for a very weak monoprotic acid can be explicitly calculated from charge and mass balance expressions:
(16)
where f is given by
while for C >> K, eq 14 reduces to , ,S
= S,, (K/Q
'/'
2112 Analytical Chemisrry, Vol. 67,No. 73,July 7, 7995
(19)
(14)
, S is linearly dependent on Cmwhere m varies from 0.5 to 1, depending on the relative magnitudes of K and C. For values of C comparable to K, e.g., put C = 2K in eq 14; this yields
, ,S
For polyprotic acids, a variety of scenarios are possible; a common one involves only the first proton being significantly, , in D2, the fully perhaps fully, dissociated in D1 (GI = G I , ~while ionized anion is the dominant species as given by eq 11. In this case, the slope Slspcan be computed by dividing eq 2 by eq 11:
f = 2[Na+lK/(K(1+ C - [Na'l)
+ K.+
The consequence of eq 20 is that f varies as a function of concentration and there is a departure from linearity in the response behavior. More importantly, for [Na+l = 0.1 mM, while an analyte of pKa = 8 is nearly completely ionized, only -10%is ionized for an analyte with pKa = 11. Clearly, detection of even weaker acids is impractical unless [Na+l is increased. Finally, while dualchannel chromatographic detection and detailed numerical processing of the resulting two channels of data is not a rare practice (see, e.g., the exemplary work of Aue et al. in ref 15 and citations therein), such data are usually collected simultaneously or in parallel. In the present case, there is a reactive element in between the two detectors that obligatorily results in time delay and dispersion. In taking the ratio of GI and Gz signals or plotting them against each other, it will be f i s t necessary to correct for this. EXPERIMENTAL SECTION
The experimental setup was similar to that used previously.12 A prototype Dionex 4OOOi gradient pump was used in combination with an auxiliary pulse dampener. Other parts, such as the injection valve (25 pL) , the electrically operated water regenerant autosuppressor (ASRS1, 4 mm), conductivity detectors (CDMI), and the knitted mixing coil, were all Dionex components (Sunnyvale, CA). All chromatography was performed with NaOH eluents, prepared with 50%NaOH and Nanopure water, at a flow rate of 1.0 mL/min. The eluent concentration varied between 3 and 30 mM; generally these were prepared in situ by dilution of a 30 mM stock. Both isocratic and gradient elution were used. An anion trap column (ATG1) was incorporated before the injection valve. The analytical column was a 4 x 250 mm AS11 column, in combination with a 4 x 50 mm AG11 guard column. All connecting tubing in the chromatographic line was 0.25 mm i.d. PEEK tubing. The MENG was made from Nafion and PTFE tubing as described below; it was powered by a previously described constantcurrent generator.12 The MENG design is an improvement over that previously used (Figure 3a, ref 12). It is constructed from Nafion 020 tubing (-0.4 mm i.d., 0.5 mm 0.d.; Perma-Pure Products, Toms River, NJ; courtesy Dr. David Leighty), having an active length of -15 mm. The electrodes are made of 100pm diameter platinum wire, of which the cathode protrudes out through the wall of the PTFE tubing. The anode is wound as a spiral around the Nafion membrane. This arrangement is inserted into an outer shell, made of a segment from a disposable plastic 1mL syringe. Feed NaOH passing through the shell was -10 mM in concentration. The NaOH solution flowed by gravity at a rate of ->1 mL/min. With a typical applied current level of 160 pA, Gbgnd typically ranged between 23 and 27 pS/cm for NaOH eluent concentrations ranging between 3 and 50 mM. The necessary voltage was 2.22.5 V. All data were acquired by a 80386based PC using Al-450 software and an ACI interface, both from Dionex. In comparing GIand Gz signals, the following algorithm was used (a) align the two peaks at the peak maximum; (b) process the leading and (15) Millier, B.;Sun,X-Y.;Aue, W. A. Anal. Chem. 1993,65,104-111.
trailing halves of the peaks separately, for GI,first compute the time interval between 25% of the peak maximum at the leading edge and the peak apex and compress the corresponding time interval for Gz to this value; (c) repeat the procedure for the trailing edge. This results in the two peaks being temporally aligned at the ho.25 points, and the data can now be directly compared. Data below 25% of the peak height are not used. The choice of h0.25 for alignment is not unique; any other value, e.g., ho.1, can be used just as well. A copy of the routine for this alignment/dispersion correction, written in BASIC, is available from the authors on request. RESULTS AND DISCUSSION Des@ and Performance of the MENG. The MENG must
be designed to minimize the induced dispersion and reduce detector noise. In addition to hydrodynamic dispersion caused by the physical nature and dimensions of the MENG conduit, there is also reactive dispersion in such a device. Eluite HX diffusing outward from the central region of the flow stream reacts with the NaOH diffusing inward from the membrane walls to form NaX. The latter is redistributed again because of the extant concentration gradient. In terms of noise, there are potentially several sources; variations of the constancy of the current fed to the MENG is generally not a significant source, but the constancy of the faradaic efficiency can be a factor since the exact physical rigidity of the device (interelectrode distance, membrane/ electrode contact., etc.) and the formation of any gas bubbles or layers on the electrode can all affect the faradaic efficiency. The trpical baseline noise (standard deviation) of the present system, operating with a Gbgnd of -25 pS/cm, is 20-25 nS/cm over a 20 min period; the value is smaller over shorter periods. On the basis of our general experience with MENG devices, the following points are salient. It is desirable to have as low a potential as possible to achieve a given current level. If the outer electrode is further away from the membrane and is, for example, in contact with the outer jacket, the interelectrode distance is typically large and the electrical resistance of the device increases. The exact physical position of the outer electrode should not be sensitive to vibrations. Both of these increase the observed detector noise. It was found that devices with an active length shorter than 10 mm tend to be more noisy than longer devices: this is understandable in view of the fact that.,at an applied current the current density is already in excess of 10 A/m2 level of 200 jA, of membrane area, a relatively high value at such low ionic strengths. At the current levels we typically use, it is interesting that devices signiiicantly longer than -16 mm also produced more noise, but this appears to be related to the relative loss of rigidity as the device length increases. Noise performance of such longer devices can be increased by improving rigidity, e.g., by coiling the inner electrode on an wettable polysulfone membrane tube before the insertion of the assembly within the N d o n membrane tube or by inserting a self-expanding helix of Nylon monofilament inside the Nafion. In any case, excessively long membrane lengths in the MENG are undesirable because they increase the hydrodynamic dispersion. Layering of gas or bubbles on the inner electrode where NaOH is generated according to the reaction Na+
+ H20+ e
-
NaOH
+ 1/2Hz
(22)
does not appear to be a significant source of noise. We studied Analytical Chemistty, Vol. 67,No. 13, July 1, 1995
2113
the use of a palladium wire cathode, which would have eliminated any evolution of hydrogen for a prolonged period of time; however, no baseline noise improvements could be discerned. Consideration of tabulated data on the aqueous solubilities of hydrogen (780 pM at 1atm and 25 "C) also indicates that there should not be any evolution of gaseous H2 accompanying the generation of 100pM NaOH. Since the aqueous solubility of 02 is much higher than that of H2, gas evolution should obviously not be a problem at the anode. Band dispersion was studied in detail for a MENG device incorporating a 15 mm long membrane; band dispersion was calculated as the square root of the difference between the square of the band volumes with and without the MENG device.'6 The dispersion values computed this way decrease with increasing band volumes; i.e., the same extent of dispersion affects a fast eluting, intrinsically narrower band more adversely than it affects a broader eluite band. For a chloride peak (injection volume 25 pL) that elutes relatively fast,the measured dispersion value was 67 f 4 ,uL when the device was operating in the passive mode (hydrodynamic dispersion only) and 94 f 6 pL with the device in actual operation. Since dispersion adds in a square root of sum of squares fashion, the contribution of the reactive dispersion can be computed to be 65 pL, i.e., it contributes as much as the hydrodynamic dispersion. Analyte Overloading. With low& acids, when C in the eluite band exceeds the background NaOH concentration in the detector, the resulting peak is W-shaped, giving the misleading impression that it is a poorly resolved doublet. This happens because the decreasing conductivity and the resulting negative signal produced according to eq 1continues only until free OHis available. When the NaOH is fully titrated, excess HX begins to increase the conductance again. If the overload is only slight, the peak may appear misshapen relative to the corresponding D1 peak. As the pKa of the eluite acid increases beyond -5, a W-shaped peak is not observed, because of the formation of a NaX/HX buffer system. At eluite PKa 2 6, after the NaOH is fully neutralized, any further amount of HX added does not dissociate significantly. Hence, analyte overload results in a flat-toppedpeak. Experimental observationsmatch theoretical expectations; simulated responses for (a) a strong acid, (b) pK = 4,(c) pK = 5, and (d) pK = 6 are shown in Figure 1. Any misshapen D2 peak should therefore be reexamined by injecting a more dilute sample. Unless overloaded, the D2 peak area/height is strictly linearly related to the injected concentration. Because the MENG responds very rapidly to a change in the driving current, the suspect D2 peak can also be reexamined in a more convenient fashion after increasing the MENG current and reinjecting the same sample without dilution. Effect of Impurities in the NaOH Eluent. Thus far, we have not taken into account the effect of impurities present in the NaOH used as the chromatographic eluent. Without an electrodialytic eluent generator (EEG)? there will always be some degree of impurities present in the eluent. Most notably, there is always some extent of C02 (or carbonate) contamination. An EEG is not commercially available; in the present work we have therefore concentrated on alternatives that would be available more easily to most users. With an impurity removing column (e.g., an anion trap column, ATC-l), it was found possible to minimize (but not completely eliminate) the level of impurities in the eluent. There (16) Dasgupta, P.K Anal. Chem., 1984,56,103-105.
2114 Analytical Chemistry, Vol. 67, No. 13, July 1, 1995
50'"' 'sb'10d'"
05 - " Time, Arbitrary Units
4 U
0
50 uM 100 pM
125
pM
150 uM
Figure 1. Responses for different concentrations (50-150 pM at the peak apex, 02 background 100pM NaOH) of a monoprotic eluite acid ranging from (a) strong acid to (d) pK = 6. The simulated peak profile is modified Gaussian and AX is assumed to be 60.
can be impurities other than carbonate in the eluent. One procedure used in this laboratory for preparing NaOH eluents involved boiling high-puritywater to remove any residual C02 and then adding high-purity solid NaOH or 50% NaOH in the desired amount. It was found that if a glass vessel is used for boiling the water and especially if the NaOH is added while the water is still hot, very substantial amounts of silicate are incorporated in the eluent. Incorporation of carbonate or silicate generally results in an increase in the eluent strength over that of pure NaOH. There is also an increase in the D1 background conductance. But the effects of these on the D1 signal are generally minor. The most important adverse effect of such impurities is their influence on the D2 signal, especially for high-pKa analytes. In the presence of significant amounts of carbonate or silicate, the absolute value of the C2bgnd is not an accurate measure of the NaOH that is free and available to react with the eluite HX. The value of & b a d does not linearly increase with the MENG current because the initial amounts of NaOH introduced are spent in titrating the impurity acids present in the suppressed background. Indeed, & b o d may actually first decrease with increasing MENG current and then increase in the expected fashion. Figure 2 shows simulated and actual Gbgnd values at two different levels of carbonate contamination. The simulations assumed a faradaic efficiency factor of 1.075, which best fits the behavior of the particular MENG device used. The close correspondence of the simulated results with the experimental data is an indication of the validity of the experimental model. Consider that, in the presence of carbonate, eq 1 will be replaced or augmented by C0:-
+ HX - HC0,- + X-
(23)
resulting in a substantial loss of signal. Further, even the occurrence of the reaction above is compromised as the pKa of HX approaches or exceeds the the pKa of HC03-. It is obvious that an approach substantially more complex than eq 10 or 11,
exponentially modfied Gaussian peak profile was assumed. The
E 0
limitingequivalent conductance of the hydrogen phthalate monoan-
1
00000Experimental Data, i=O G,,,=0.29 uS/cm - Simulation, 1.9 u M H2C03, GmW=0.29 S/cm MAMExperimental Data, i=O GI -1.0 uS/m Simulation, 17 uM H2C03, "e",-,=l.O US/cm
______
0 0
Figure 2. Simulated (lines) and experimental (points) for the second detector background conductance at two different carbonate contamination levels. The carbonate concentration used in the simulation was computed from the first detector background conductance, assuming it to be solely due to HzC03.
-7.0
11
:W'
\;/
I
I
I I
I
I
I I
,
1
Experimental r r
I
I
,
I
I
I
I
I
-7.0
Simulated
I
Time Figure 3. Dependence of the D2 analyte response on the D2 background conductance in the presence of significant eluent impurity levels. (a) Experimental and (b) theoretically simulated results are shown for 100 pM phthalate as sample at two different carbonate contamination levels (at two different 0 2 background conductance values for the higher contamination level).
that takes into account the presence of the various carbonate species and their redistribution upon reaction with HX, is necessary to appropriately simulate the system. Figure 3a shows the experimentalresults for 100 pM phthalate as the injected sample and 30 mM NaOH as the eluent. The presence or absence of an impurity trap column made a significant difference in the D1 background conductance (0.88 vs 1.64 pS/cm). The results at two different MENG current levels (to obtain & b a d values of -15 and 25 pS/cm) for the higher carbonate contamination level and for a single MENG current level (GZbgnd -15 pS/cm) at the lower carbonate contamination level are presented. A numerical simulation of the same system is shown in Figure 3b. The carbonate contamination levels were calculated to be 13.3 and 42.8 pM, respectively, with and without the impurity trap column, assuming that the D1 background conductance is solely due to HzC03. An
ion and the phthalate dianion are not available in the literature and were respectively assumed to be 29 and 57 pS/cm. The exact peak width is different in the experimental vs the simulated system; also, experimental detector calibration may not have been exact. With this in mind, the agreement between theory and experiment is excellent. Since the presence of carbonate in D2 is detrimental, we wished to determine whether the choice of the MENG feed solution has any effect on the system performance. It was thought that the choice of a neutral salt solution such as 10 mM Na2S04 instead of the NaOH as the Na+ feed solution may reduce the amount of carbonate that is introduced through the MENG, if any is introduced through this route. Analyte peak heights were unchanged whether NaOH or Na2S04 was used as the MENG feed. The use of Na2S04 or similar other salt solution, however, electrodialyticallygenerates acid on the feed side, and the H+ thus generated competes with Na+ for passage to the cathode side . (very effectively, since LH is 7 times greater than L N ~ The electrodialytic NaOH generation efficiency is thus markedly decreased relative to the use of a NaOH feed. Consequently, a higher MENG current is needed to attain the same Gbgnd with a concomitant rise in the noise level. Therefore, the use of NaOH as the Na+ feed is preferred. As long as electrodialytically generated NaOH is unavailable, we find that minimum carbonate contamination is achieved by diluting a more concentrated stock eluent solution (maintained in a bottle with a soda lime-based COZtrap) in line with highpurity water if a gradient pump that allows in situ mixing is available. Typically 10-20% of the stock eluent should be diluted with 8 0 4 0 % water; satisfactory performance is then obtained in the presence of an impurity trapping column. The presence of the latter is still critical, however. As a general rule, optimum performance can be expected from the system as long as the D1 background conductance is 5 1 pS/cm. System Noise and Iimits Of Detection. The limits of detection for various anions at the second detector, based on a signal equal to 3 times the standard deviation of the background signal, is listed below (in micromolar, followed by retention time in minutes in parentheses): azide, 3.0 (4.58); arsenite, 3.2 (2.85); borate, 8.6 (3.65); bromate, 1.65 (3.63); bromide, 1.7 (4.25); chloride, 1.5 (2.48); malate, 1.9 (7.22); nitrate, 2.7 (5.98); nitrite, 2.1 (3.28); phthalate, 1.2 (5.03); silicate, 4.0 (2.68); succinate, 1.7 (7.00); sulfate, 2.1 (8.08), and sulfide, 5.1 (3.03). There is some indication that thermostating the entire setup results in significant further improvement of signal/noise; detailed results will be published in the future. Borate and cyanide peaks show significant tailing and broadening on the stationary phase used; this adversely affects the attainable limit of detection. For volatile weak acids such as HCN and HzS, transmembrane loss in the suppressor can also 0ccur.17 Calibration Behavior. Second detector calibration shows excellent linearity for a variety of anions tested across the pKa range. Coefficient of determination (linear 72 , peak area vs injected concentration at a &bgnd level of 25-27 pS/cm) values for arsenite, chloride, monochloroacetate,nitrate, nitrite, phthalate, silicate, succinate, and sulfate were respectively determined to (17) Dasgupta, P. IC J. Chromatop. Sci. 1989,27,422-448.
Analytical Chemistry, Vol. 67, No. 13,July 1, 1995
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be 1.oo00, 0.9999, 1.0000, 0.9999, 0.9999, 1.oo00, 0.9998, 0,9999, and 1.0000, over the 0-500 pN injected sample concentration range. Lhearity of calibration for several of these analytes is much worse on the first detector; some are not even seen by the fist detector. The response linearity observed for silicate is slightly worse because at the high end of the injected sample concentration the extent of ionization for silicate at the second detector is concentration dependent, decreasing at higher concentrations, as elaborated in eqs 20 and 21. Figure 4 shows a sensitivity plot as a function of concentration. Essentially uniform sensitivityis seen for all the analytes tested (with the recognition that at low concentrations,blank correction becomes increasingly important) with the exception of arsenite. In the arsenite case, oxidation to arsenate, either on-column or in the prepared standards, to a relatively greater extent at lower sample concentrations, is the likely reason. The calibration slopes for the various analytes in the above experiment are shown plotted in Figure 5 against relative theoretical sensitivities obtained from eqs 10, 11, 20, and 21. The theoretical values were calculated based on an assumed injected concentration of 125 pM,a 10-fold dilution (at the peak apex) during the chromatographic process, a D2 background of 100pM NaOH, pKa values of the acids, and the limiting equivalent ionic conductance values. The latter data are not available in the literature for phthalate, silicate, and arsenite. These values were initially estimated from their size and shape with later experimental verification by ion-exchange/conductance measurement techniques as detailed in ref 18. The values used were 67, 44.5, and 57 pS/cm for the arsenite and silicate monoanions and the phthalate dianion, respectively. The pKa value(s) and any a s sumption regarding dilution in the chromatographic process are needed only for the very weak acids because of the potentially nonlinear relationship of the signal with concentration represented by eqs 20 and 21. The linear relationship observed between theoretical predictions and experimental data suggests that if pKa and equivalent (18)Dasgupta, P.IC; Nara, 0.Anal. Chem. 1990,62, 1117-1122.
21 16 Analytical Chemistry, Vol. 67, NO. 13, July 7, 1995
conductance values are known, reasonably good results can be obtained without actual calibration with analyte standards. This can be a real advantage where standards are difficult to prepare, expensive, or degrade rapidly, e.g., by oxidation. It is important to note that the best-fit line in F i i e 5 was drawn without considering the data for arsenite, silicate, and phthalate for which the equivalent conductance values were not tabulated. It is remarkable as to how well the predicted values in these cases fall on the best-fit line. Nevertheless, the approach should be implemented with caution for the anions of very weak acids where the pKa values are comparable to or greater than the pH of the D2 background. Not only is it necessary to use the correct equivalent conductance value but the results can be particularly sensitive to the choice of the pKa value. In a significant number of cases, the reported pKavalue spans a significantrange; Perrin's compilation of pKa values19 show, for example, the pKa value for HAsOz at 25 "C to range from 9.08 to 9.28. There are other cases where the tabulated equivalent conductance value may bear reexamination. As Figure 5 shows, nitrate and nitrite have tabulated equivalent conductance values that are almost the same but have perceptibly different experimental calibration slopes. It is interesting that we have previously observed, in a totally disparate experiment, that there is also a discrepancy between the behavior of nitrite and nitrate when the electrophoretic mobilities are plotted against the tabulated equivalent conductance.20 Estimation of Iimiting Equivalent Conductance. The present experimental setup permits the estimation of the equivalent conductance in a unique manner if the sample concentration in equivalents per liter is known (a knowledge of the relevant PKa value($ is also necessary if very weak acids are involved). Equations 10,11,and 20 indicate that the peak area is proportional to the injected concentration C (in equiv/l; for very weak acids the fraction ionizedfcan be computed from eq 21) and the difference in the equivalent conductance between the hydroxide ion and the test analyte. Consider eq 10; if the area response values for different analyte ions at the same concentration are plotted (19)Pemn. D.D.Ionisation Constants of Inorganic Acids and Bases in Aqueous Solution; 2nd 4.;Pergamon: New York, 1986. (20)Dasgupta, P. IC; Bao, L Anal. Chem. 1993,65, 1003-1011.
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