802
Anal. Chem. 1987, 59, 802-808
Gradient Anion Chromatography with Hydroxide and Carbonate Eluents Using Simultaneous Conductivity and pH Detection Hideharu Shintani' and P u r n e n d u K. Dasgupta* Department of Chemistry, Texas Tech linipersity, Lubbock, Texas 79409-4260
High exchange capacity membrane suppressors make gradient anion chromatography practical, with submicromolar detection limits attalnable for a number of common anions when a data ecqulsition system is used to store a blank run and perform background subtraction. Wllh hydroxide eluents, pH measurement yields attractive detectlon ilmits. Postsuppressors, based on a porous polypropylene tubular membrane as is, or coated with silicone rubber, permit acceptable performance with the more commonly used carbonate-based eluents. For isocratic elution wlth hydroxide eluents, direct potentiometrlc pH detectlon allows limits of detection only slightly worse than conductometry and Is adequate for most purposes.
While conductometric anion chromatography with isocratic elution is a mature technique with hundreds of papers on fundamental methods and applications already published ( I ) , gradient ion chromatography (IC) with conductometric detection is still largely in its infancy. The limited number of reports that have appeared to date on gradient conductometric IC all deal with suppressed systems; barring a major scientific breakthrough, it is not possible to adapt gradient elution to single-column conductometric IC. The first report on gradient conductometric IC, utilizing carbonate eluents, was largely exploratory (2). Nevertheless, it served to establish that pitfalls abound in approaches involving a gradient between identical molar concentrations of bicarbonate and carbonate, which may simplistically be assumed to lead to the same carbonic acid concentration after suppression. A subsequent report by the same Swedish researchers attempted to make the technique more practical by incorporating a porous PTFE membrane-based postsuppressor to remove carbon dioxide from the suppressed effluent ( 3 ) . While the concept was demonstrated, large band broadening and frequent leakage remained problems. Thin silicone rubber tubular membranes immersed in warm alkali permitted permeative removal of C 0 2 and represented a more practical approach (4). Postsuppressors are essential to perform gradient IC at trace levels with carbonate eluents; Tarter's work ( 5 ) has shown that while one may perform higher level determinations without such devices, the prospect is rather limited. A different approach was taken by one of us ( 6 ) using a nonpolar macroreticular poly(styrenediviny1benzene) column and running a gradient between tetrapropylammonium hydroxide (which provides retaining power via ion interaction, see ref 1) and sodium hydroxide (which provides eluting power), in conjunction with a filament-filled helical (FFH) membrane suppressor. Limited exchange capacity of the first-generation FFH suppressor for large cations like NPr,+, as well as impurities in the eluent, proved to be the primary limitations. More recently, Irgum ( 7 ) has introduced amil Permanent address: Department of Medical Devices, National Institute of Hygienic Sciences, Tokyo, Japan.
0003-2700/87/0359-0802$01.50/0
noalkylsulfonic acids as eluents for this application and has shown the power of compound gradients with a poly(methacrylate) stationary phase. Along with the introduction of their gradient IC instrumentation, researchers from the Dionex Corporation have presented their work on gradient IC ( 4 9 ) . The present work demonstrates the utility of suppressed hydroxide eluent gradient IC. EXPERIMENTAL SECTION The prototype version of a Model 4000i gradient ion chromatograph (Dionex Corp., Sunnyvale, CA) equipped with an anion micromembrane suppressor (AMMS) and a conductivity detector was used for all work. A flow-through cell equipped with a flat surface pH electrode (Markson Science, Phoenix, AZ) connected to a Phi 71 pH meter (Beckman Instruments, Fullerton, CA) was used in series following the conductivity detector. A homemade stainless steel replica of the Plexiglas flow cell supplied by Markson was used for pH measurement, with the cell body grounded to the pH meter. The estimated volume of the flow cell is 10 pL. (The Plexiglas cell is more susceptible to noise induced by external electrical fields than its metallic counterpart and is not recommended.) Both conductivity and pH outputs were recorded simultaneously on a dual channel strip-chart recorder (Knauer, TY-2, Sonntek Inc., Woodcliff Lake, NJ) and the data were also concurrently acquired by a Chromatopack C-R3A data acquisition system (Shimadzu Scientific, Columbia, MD) for postprocessing. Only a minor modification was made to the Dionex prototype instrument: A single bead string reactor (SBSR, see ref 10) made from a 17 cm length of a 1.5 mm i.d, 3 mm 0.d. PTFE tube filled with 1 mm diameter glass beads (with glass wool plugs at each end to retain the beads) was substituted for the existing line connecting the pump intake to the solenoid-controlled eluent manifold. Past the SBSR, 0.8 mm i.d., 1.5 mm 0.d. tubes were used up to the inlet check valves. These modifications resulted in significantly better mixing and reduced the holdup volume by more than 50%, thus reducing the time between the programmed gradient and its actual appearance at the column inlet. The current commercial version of this instrument incorporates a second static mixer on the high-pressure side and reportedly reduces pump noise further-this modification was not in place at the time these experiments were conducted. Unless otherwise noted, chromatography was conducted on a Dionex AS4A column (4.0 X 250 mm) without a guard column, at an eluent flow rate of 1 mL/min and a sample volume of 50 PL. Hydroxide eluent solutions were prepared by adding calculated amounts of concentrated (50% (w/v)) stock carbonate-free hydroxide solutions (highest commercial purity available) to C02-free water (boiled for 22 h and transferred hot into a collapsible plastic bag). The eluent reservoirs were pressurized at 3-5 psi from a compressed house air source, transmitted through soda-lime tubes. Actual hydroxide concentrations in the eluent(s) were determined by withdrawing aliquots through the pumping system and performing acidimetric titrations with primary standard potassium acid phthalate. Carbonate eluent strengths are based on the weighed amounts of the pure dry compounds. Sulfuric acid (12.5 mM) was used as the regenerant for the AMMS and delivered from a reservoir pneumatically pressurized at 15 psi. Construction of the Membrane Postsuppressors. Microporous tubular polypropylene membranes (400 pm id., 25 pm wall, nominal pore size 0.02 pm, surface porosity 4070, Celegard X-20, Celanese Corp., Charlotte, NC) were coated with silicone rubber by immersing a roll of the membrane tube in a 3 4 % (w/v) C 1987 American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 59, NO. 6, MARCH 15, 1987 803 solution of silicone rubber adhesive (General Electric, translucent type for household use) in dichloromethane. The membrane was allowed to sit in the solution for a minute and then the solution was decanted off and the membrane tube was uncoiled and suspended from a suitable spot and allowed to dry (in a dust-free environment) for 2 2 h. The membrane was recoated repeatedly in this fashion, up to 10 times, until the coating was deemed to be leak-free. Testing for pinhole leaks was conducted by injecting methanol into the lumen of the fiber with a 27-gauge hypodermic needle (which fits tightly in the fiber) while the other end of the fiber is pinched shut manually. With reasonable manual pressure on a 1-mL capacity syringe (this amounts to about 50-100 psi), no methanol should appear through the fiber walls when the coating is leak-free. Note that water cannot be substituted for methanol because water does not wet the hydrophobic membrane and it requires considerably greater pressure to initially establish water flow across the membrane, even for the virgin membrane tube. The coated membrane tube was cut into 30-cm sections and 250 pm diameter nylon monofilament (10 l b strength fishing line, STREN, Du Pont) was inserted into each of the hollow fiber sections, with lengths of the filament protruding out of each end of the hollow fiber. Typically, 12 such sections were taken and wrapped as a uniform parallel ribbon on a 1.5 mm diameter stainless steel support rod. The ends were affixed to the support with adhesive tape, leaving -2 cm free at each terminal end. The assembly was then immersed in boiling water for 30 min to thermoset the filament, similar to the FFH Nafion membrane suppressors described previously (11). After the assembly was thermoset, the adhesive tapes and the filament-filledmultistrand helical tube assembly were removed from the support. The terminal ends were inserted into hex-head 1/4-28 threaded nuts, the nut heads facing each other. A few millimeters of the fiber ribbon was allowed to protrude out of the sealing sides of the nuts. Polyurethane spray foam was squeezed in from the head side of the nuts with a fine nozzle until it appeared on the sealing sides. The polyurethane was allowed to cure and excess foam and protruding membrane were cut off flush with the sealing surface of the nuts with a sharp razor blade, revealing multiple openings of the filament-filled fiber bundle. The assembly was then enclosed in a Tygon jacket tube, provided with inlet/out tubes for a 50 mM NaOH solution (to act as sink for the COz removal) and sealed with polyurethane foam. The sodium hydroxide flow was maintained by gravity, at 1.5 mL/min. Membrane postsuppressors based on the virgin Celgard membranes were made in a similar fashion, except 240 pm i.d. hollow fibers filled with 4 lb strength fishing line (200 pm diameter) were utilized. This device was operated with COz-free air (house air bubbled through 5 M NaOH) passing through the outer jacket. A third type of membrane postsuppressor was based on the report of Siemer and Johnson (4). Silicone rubber tubes (250 pm i.d., 100 pm wall; Patter Products, Beaverton, MI) were used in the form of a 10-fiber bundle of 30 cm length with silicone rubber adhesive providing the scaling inside the nuts. This device was operated with 50 mM NaOH flowing in the external jacket. Gradient ChromatographyConditions. The elution conditions used for the gradient chromatograms shown are as follows: Figures 5, 7 , 8: eluent 1, H20; eluent 2, 204 mM KOH. Increases in % 2 between specified time points
-
time 0
-- - 15
8
25
38
% 20+3-+10-20-55
Figures 6, 9: eluent 1, H20; eluent 2, 50 mM NaHCO,; eluent 3, 50 mM NaZCO3. Gradient program was time 0
-- - - 7
step
7.1
%25-5-
0
%30-+0+
3-
13
24
30
13-+30+50
RESULTS AND DISCUSSION Effect of the Cation on the Suppressing Ability of the Membrane Suppressor. While the choice of the cation counterion in the eluents used for anion chromat,ography (e.g., LiOH vs. NaOH vs. KOH) does not have any effect on elution
6
100
150
50
100
1
[M+], m M
carbonate
Flgure 1. Exchange capacity of the AMMS with hydroxide and carbonate eluents, 12.5 mM H2S04regenerant at 15 psi: (1) Li,CO,; (2) Na2C03;(3) K,C03; (4) LiOH; (5)NaOH; (6) KOH. The noise level is depicted with each trace; the shaded area indicates the noise amplitude. Note that the origin begins at 50 mM for hydroxide eluents.
behavior, the maximum attainable dynamic suppression capacity in any given membrane-based suppressor is dependent on such a choice. For a hydrodynamically well designed device such as the AMMS (12),it is not likely that large differences exist in rates of transport to the membrane among various cations. Considerable turbulence must exist in such a device due to the presence of ion exchange screens in the flow path and transport to the membrane cannot be diffusion-dominated. Indeed, if the overall suppression capacity in the AMMS were to be determined by the rate of ion transport to the membrane, no differences in the order of suppression capacities are expected in going from hydroxide to carbonate based eluents. Experimentally, however, the order does vary (vide infra). The suppression capacity is likely dominantly controlled by two factors: (a) the uptake of the cation by the membrane-this is determined in turn by the selectivity coefficient of the membrane for the cation and the presence of competing ions, namely, H+; (b) the rate of transport of the cation through the membrane. A detailed mathematical description can be found in ref 11. It is only for hydroxide based eluents that H+ concentration remains negligible throughout the entire exchange process and thus factor (a) has little effect-membrane sites are automatically saturated with the eluent cation. The AMMS device used in our study was able to quantitatively suppress 155 mM KOH, 140 mM NaOH, and 105 mM LiOH, respectively, all at a flow rate of 1 mL/min. The quantitative aspect of the exchange is best determined from the absolute noise level in the conductance detector. In Figure 1, the shaded region for each curve indicates the onset of significant detector noise; the amplitude of the shaded area is the noise amplitude. The abrupt rise in noise levels past the maximum capacity of the suppressor is evident. Note that membrane-based devices vary considerably in performance-the above exchange capacities may or may not represent the performance of a typical AMMS device. Further, under the test conditions, the exchange capacity is limited by the availability of regenerant. It is possible to increase the stated exchange capacities further,
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ANALYTICAL CHEMISTRY, VOL. 59, NO. 6 , MARCH 15, 1987
within limits, by increasing regenerant concentration and/or flow rate With a carbonate eluent, as ion exchange progresses, H2C03 accumulates and the competition of H+ thus generated for the ion exchange sites becomes significant. As discussed in more detail elsewhere (13),maximum suppression capacities attained with hydroxide eluents cannot, for this reason, be attained with carbonates or other salts. Also, because of the existence of significant concentrations of H+, membrane sites are not saturated with the metal ion and the selectivity factor plays a role. The AMMS device used in our study was found to have a suppression capacity of 40 mM K2C03, 45 mM Na2C03,and 35 mM Li,CO,; note that not only is the order different but all of these capacities are significantly lower than the corresponding values for the hydroxide eluents. pH Detection. In suppressed anion chromatography, the suppressed eluent pH is presumably near-neutral to only slightly acidic, because only a salt of weak acid (including water in this consideration as a Bransted acid) can be used as an eluent. The same phenomenon that makes conductometric detection of strong acid anions particularly attractive in suppressed IC is expected to lead to successful pH detection. The elution of the sample anion is manifested by the appearance of a fully dissociated acid, which is accompanied not only by an increase in conductivity but also by a concomitant drop in pH. In the past, such changes in pH have been measured indirectly by the postcolumn introduction of some electroactive species which participates in a redox equilibria involving the hydrogen ion (for example, introduction of quinone and the quinone-proton-hydroquinone equilibria) and using an electrochemical detector ( 1 4 , 1 5 ) . In fact, even without the intentional introduction of any electroactive species, the background current of electrochemical detectors with aqueous eluents is acutely pH dependent in the absence of large amounts of a supporting electrolyte and increases with decreasing pH. Although it has not been clearly identified as pH detection, Tarter's observations (5, 16) that an electrochemical detector placed after the suppressor responds to anions that are not electroactive are thus readily explicable. While the direct potentiometric sensing of pH may not be the most sensitive detection technique, it is difficult to understand the total lack of attempts at such detection in view of the facts that a pH meter is one of the most common fixtures in an analytical laboratory and a host of potentiometric methods to detect eluting anions using ion-selective electrodes have appeared in the literature ( 1 ) . Note also that some recorders and data acquisition systems in use today have sufficiently high input impedances to permit direct connection with the pH electrode without the intervention of a pH meter. Further, Ruzicka and Hansen (17) have clearly established for FIA applications of ion selective electrodes that an inclined electrode and gravity effectively produce a "flow cell" of 10 pL volume and an enclosed flow-through cell is really not necessary. Hydroxide eluents, which produce the highest background pH, should be particularly amenable to pH detection. Consider that with a typical carbonate-based eluent, 2.4 mM Na2C03+ 3 mM NaHC03, the detector sees 5.4 mM H2C03 as background. With 6.3 as the pK1 for H2C03(18), [H+] is 52 pM and pH is 4.28. When an analyte band, composed of 1.5 ppm (42 HM) C1- a t the peak, elutes, the effluent composition a t the peak is 42 WMHC1, 5.38 mM H2CO3. From mass and charge balance considerations it is easily computed that the peak composition is [Cl-] = 42 WM,[HC03-] = 35 WM, and [H'] = 77 pM (pH 4.11). The net signal (ApH) is therefore 0.17 pH units. In contrast, for an hydroxide eluent which is suppressed to water, background pH (hypothetically) is 7 and 42 pM HC1 in the effluent peak means a pH of 4.38
-
0.4 5 0
~
.I
-
0.375
L2
lp
4 -,0.225,
-
I
v)
E 0,
-c 0.150-
I
20
40
60 80100 200 Anion c o n c e n t r a t i o n , m
Figure 2. Calibration plots for three common anions in pH detection under isocratic elution conditions: s o l i lines, 30 mM KOH eluent: dashed lines, 5 mM Na,C03 eluent; (1) CI-, (2) S O:-, (3) NO3-.
and a net signal of 2.62 pH units results. Even more importantly, the relative difference between the two eluent systems increases with decreasing analyte concentration. Admittedly, however, a pH of 7.0 is difficult to attain in practice even for a hydroxide eluent. This is due to impurities in the eluent, including intrusion of atmospheric C02, and acidic regenerant penetration through a membrane suppressor (19). While detection limits are certainly better for hydroxide rather than for carbonate eluents when using pH measurements, we are presently unable to fully elucidate the phenomena that govern the absolute value of the base line pH sensed by the potentiometer for suppressed hydroxide eluents. The base line pH with suppressed carbonate eluents are very close (within 0.1 pH unit) to theoretically calculated pH values (4.65, 4.50, 4.30, 4.15, 4.00, and 3.80, respectively, for 1, 2, 5, 10,20, and 50 mM carbonate). The hydroxide eluents lead to essentially pure water and the actual value of the base line pH is dependent on the choice of the cell construction material and the electrode. With in situ two-point calibration with pH 4 and 7 standard buffers, the typical base line pH with 10-100 mM hydroxide eluents reads between 4 and 5 . From the conductivities measured immediately prior to the pH cell, it is clear that these solutions cannot possibly contain lo4 M H+. Any significant contribution of COP can be ruled out because the placement of a postsuppressor device for C 0 2 removal between the conductivity and the pH detectors has no significant effect on the value of the base line pH. Measurement of sulfate in the detector effluent also confirms that the penetration of regenerant is negligible under our experimental conditions. It is likely that the high resistivity of the medium leads to an aberrant base line pH readout and streaming potential effects may be significant. However, the real dilemma is in the fact that not only is the base line pH quite stable, the response behavior is consistent with the apparently registered value of the base line pH. This is illustrated by the following. The pH response data for a 30 mM KOH eluent and a 5 mM Na2C03eluent on a AS4A column for three common ions, chloride, sulfate, and nitrate, are shown in Figure 2. The concentration axis is logarithmic and the pronounced asymptotic nature of the response is theoretically expected for an eluent containing a finite amount of background acidity. Consider that if the background pH is x (the acidity being completely contributed by penetration of a strong acid regenerant), the background [H'] is lo-". If the sample anion (monovalent) concentration at the peak apex is y M, [H+] a t
ANALYTICAL CHEMISTRY, VOL. 59, NO. 6, MARCH 15, 1987
805
Table I. Conductivity vs. pH Detection: Limits of Detection: Isocratic Elution, Hydroxide and Carbonate Eluents eluent 30 m M KOH conductianalyte
tR, min
vity
pH
chloride nitrite nitrate sulfate
2.3 2.8 4.5 5.5
0.08 0.25 0.30 0.10
0.08 2.0 2.0 1.0
eluent 2.5 m M Na2C03 conductit ~min , vity pH 2.2 2.7 5.1 10.5
0.15 0.40 0.50 0.40
/cI'
5.0 6.0 6.0 5.0
poi-
aIn r M units, S I N = 3, Dionex AS4A column, flow rate 1 mL/ min.
+
the peak apex is lo-" + y and the pH is -[log (10-x y ) ] . The net signal, z, in ApH units is therefore z =x
+ log (10- + y )
(1)
Algebraic rearrangement of eq 1 leads to the form
loz = 1 + l0Xy
-
0
(2)
which is easily plotted; We can treat the data in Figure 2, for example, those for chloride, in this manner. From the injected concentration, the peak concentration can be approximated from the base width of the peak and assuming that the concentration at the peak is twice that obtained by uniform dilution of the sample volume (50 pL) in the volume represented by the base width of the peak (350 pL). A plot of IO2 vs. y for the chloride data in Figure 2 is linear with a correlation coefficient better than 0.9998. Further, the intercept is 1.023, in excellent agreement with theory and the value of the base line pH, x , is obtained from the slope to be 4.4,essentially identical with the base line pH value registered by the potentiometer. We made limited attempts to carry out substoichiometric suppression by restricting regenerant flow such that the base line pH will read between 6 and 8. Not only was the base line pH relatively unstable as might be expected, but the net signal for any given sample concentration also decreased. Introducing a base postcolumn through the membrane reactor (see, for example, ref 20) after regular suppression may be a more viable approach to understand the behavior of the system but was not explored in the present study. It is clear, however, that the detection limits offered by direct potentiometric pH detection with suppressed hydroxide eluents are adequate for many applications. The limits of detection for four common anions are shown in Table I for conductometric and direct pH detection with both hydroxide and carbonate eluents under isocratic elution conditions. Typical sample chromatograms with a isocratic hydroxide eluent are shown in Figure 3. Figure 4 shows the separation of a more complex mixture with an isocratic bicarbonate eluent. If C 0 2 is removed with a silicone-coated membrane postsuppressor, the linearity of calibration at low level is better, but the gain in attainable limits of detection for conductivity detection is marginal; the without postsuppressor chromatogram is therefore not shown. The absolute value of the background conductance, however, decreases considerably with the postsuppressors, up to 85% depending on the device and the eluent concentration. It is interesting to note that no significant decrease was observed in the background conductance when postsuppressors were used with hydroxide eluents indicating that the latter were indeed relatively free from COz contamination. For carbonate eluents, the increase in sensitivity for pH detection upon COzremoval is much more pronounced, as shown in Figure 4 (A vs. B). However, the significant volume of the postsuppressor deteriorates resolution in either detection mode. Note that aside from any broadening due to the connecting line between the conduc-
5 10 Minutes
Flgure 3, Isocratic hydroxide elution (30 mM KOH) chromatogram of three common sample ions (5 nmol each) with pH and conductivity detection.
0
10
5
C Condurtivity detection after
cop ,.rn.Y.J 7
6
L
0
I
I
5 10 Minutes
Flgure 4. Isocratlc carbonate elution (2.5 mM NaHCO,) chromato(2)CH,COO-; (3)HCOO- ("$X&gram. Peak identification: (1) IO3-; coelutes); (4) BrO,-; (5) CI-; (6) NO?-; (7) N3-.
tivity and the pH detector, the logarithmic nature of pH response results in an apparent perception of decreased resolution-this is merely an artifact of a logarithmically dependent ordinate response. Gradient Elution. As a general rule, linear, rather than step gradients cause less base line disturbance for either hydroxide or carbonate eluents. Figure 5 shows the separation and detection of a large number of anions with gradient hydroxide elution and simultaneous conductivity and pH detection. Figure 6 shows the analogous case with a carbonate eluent. Although some improvement results with postsuppressorbased C02removal, the situation is still far less satisfactory compared with gradient hydroxide elution. Attainable detection limits under gradient elution conditions are shown for four common anions in Table 11. Gradient Hydroxide Elution: Eluent Precolumns and Background Correction. The fundamental problem in carrying out trace analysis under gradient elution conditions is that the impurity levels in eluents begin to approach the analyte levels of interest and, more importantly, these im-
806
ANALYTICAL CHEMISTRY, VOL. 59,
NO. 6, MARCH 15, 1987
17
'- , V
I
I
I pH
CO?
removal
/'
-1
-
.
1
15
20
25
30
40
35
Minutes
Figure 5. Simultaneous pH and conductivity detection with gradient
hydroxide elution. See experimental section for gradient program. Peak identification: (1) iodate; (2, 3) fluoride and acetate; (4) formate; (5)sulfamate; (6) bromate; (7) chloride: (8) nitrite; (9) nitrate (azide coelutes here): (10)sulfate: (11) malate; (12) tartrate; (13)oxalate: (14) phthalate; (15) chromate: (16) phosphate: (17) citrate. Each anion concentration is 100 KM. Table 11. Gradient Hydroxide and Carbonate Elutions: Limits of Detection" with Conductivity and pH Detection analyte
chloride nitrite nitrate sulfate
hydroxide gradientb conductivity pH 0.1 0.9 0.9 0.1
carbonate gradient? conductivity pH
0.4 2.0 2.0 1.0
0.2 0.5 0.5 0.5
_
_
I
_
_ L - _ d ~ _ L _ _ _ 1 - - .
i
10
5
20
15
25
Minutes
Figure 6. Gradient carbonate elution, pH, and conductivity detection with and without postsuppression. See Experimental Section for gradient program. Peak identification: (1) iodate (fluoride coelutes); (2) acetate; (3)formate (sulfamate coelutes): (4) bromate; (5) chloride; (6)nitrite; (7) azide; (8) nitrate (chlorate coelutes);(9) sulfate (sulfide, phosphate coelute);(10) malate (tartrate coelutes);(1 1) oxalate; (12) phthalate: (13) chromate: (14) citrate.
5.0 8.0 8.0 5.0
In pM units, S / N = 3, Dionex ASQA column, flow rate 1 mL/ min. *Gradient program is the same as that in Figure 5 , background correction applied, no cleanup column. Gradient program is the same as that in Figure 6, no postsuppressor, background correction applied. _
-
0
5
0
15
10
20
25
30
35
40
~
purities concentrate on the analytical column during the initial portion of the run and are later eluted as the eluent strength increases. For this reason, the use of a high capacity strong base exchanger in the eluent ion form between the pump and the injector has been advocated (6) since the early days of gradient conductometric IC. Favorable experiences with such approaches have been reported more recently (9). Therefore, we experimented with the placement of a 250 X 4.6 mm precolumn filled with Dowex-2x8 in hydroxide form (200 mesh, the column must be washed with strong carbonate-free hydroxide for prolonged periods to completely remove all traces of chloride (in which form the resin is supplied) before the injector. The result is shown in Figure 7 . The rise in background is decidedly reduced makmg accurate quantitation of nitrate (peak 9) possible. The change in eluent strength due to removal of impurity ions is also marked-the last peak (peak 17, citrate) exhibits almost a 5-min increase in retention time in the presence of the Dowex-2x8 column. The change in eluent strength is also presumably the reason for deterioration of resolution between peaks 11, 12, and 13 in the presence of the precolumn. However, this resolution can doubtless be restored utilizing a different gradient program.
t--. 0
1
5
10
15
I
I
I
20
25
30
>-
35
, 40
Minutes Figure 7. Improvement in base line perturbation with the use of an eluent cleanup precolumn in hydroxide gradient elution: A, with pre-
column; 8, without precolumn. Chromatographic conditions and peak identities are as in Figure 5.
A superior but more expensive alternative in our judgment to eluent cleanup columns is background correction. Figure 8 shows uncorrected and corrected chromatograms with a AS5 column, reportedly designed specifically for use with hydroxide eluents. A blank chromatogram (obtained with the same gradient program without sample injection) was stored and subtracted from the uncorrected run to perform the base line correction. The resolution of early eluting ions is decidedly better on this stationary phase and a significant decrease in retention time for late eluting ions, compared to the AS4A column, is attainable without sacrificing resolution. We regard
ANALYTICAL CHEMISTRY, VOL. 59, NO. 6, MARCH 15, 1987
807
nj2
C
L
0
8
5
1
10
I
15
I
20 25 M i n Utes
I
30
Figure 8. Applicability of base line correction In gradient hydroxide elution. AS5 column; other conditions as given in Figure 5.
the background correction alternative to be superior for two reasons. First, the blank gradient background appears to be highly reproducible over a significant period of time, obviating the necessity of having to run a blank with every sample run. Second and perhaps more importantly, the impurity ions captured by the cleanup column do eventually elute and may do so at the least opportune time, completely rendering a valuable, even irreplaceable, sample run worthless. Gradient Carbonate Elution: Postsuppressors and Background Correction. Practical carbonate gradient programs involve initially bicarbonate and a final gradient program in carbonate (see, for example, Figure 6). The resulting change in charge and concentration of the eluent ion during the run obviates any beneficial use of a cleanup column; the situation has been described previously in adequate detail (2). In the absence of a data acquisition device, some sort of a postsuppressor device to remove CO, is essential to keep the attainable limits of detection reasonably attractive. The single fiber silicone rubber tube postsuppressor operated at elevated temperatures ( 4 ) is woefully inadequate at the high carbonate eluent concentrations (during the final portion of the gradient run) used in this work. Further, in the absence of active thermostating, we find that the acute temperature dependence of conductivity leads to sufficiently increased drift and decreased precision such that little or no gain results in attainable detectabilities. A multiple strand silicone-rubber-tube-based postsuppressor was constructed but performance was distinctly poor compared to the alternatives below. Not surprisingly, CO, transport rate is by far the best with uncoated porous membranes and such a postsuppressor shows the best results, although quantitative removal of COz is still not achieved. Note that part of the rise in background is due to impurity ions in the eluent and not due to increasing COP content of the suppressor effluent. However, it is common for uncoated membrane tubes to develop leakage upon extended operation (this is minimized by using COz-free air as the external sink rather than an alkali solution, but not completely eliminated). The silicone-rubber-coated porous membrane postsuppressor was developed for this reason and represents a compromise between leakage problems and efficient COP removal. The results with and without these postsuppressors are shown in Figure 9. Additionally, Figure
I
I
I
0
5
10
I
/
20 Minutes 15
I
1
25
30
Figure 9. Postsuppressionvs. background correction in gradient elution with carbonate eluents: (A) chromatogram without postsuppression or base line correction; (B) with the use of silicone rubber coated postsuppressor; (C) with the use of porous membrane postsuppressor; (D) base line correction performed on A. Chromatographic conditions and peak identities are given in Figure 6.
9 shows background correction performed on the no postsuppressor chromatogram and this is clearly the superior alternative. The relatively large amounts of membrane required to effect significant CO, removal represent a significant dead volume-the deterioration of resolution of the early eluting ions is noticeable with the use of the postsuppressors. At the present time, the continued prospect of using carbonate-based eluents for gradient IC seems rather limited. The only advantage of carbonate eluents is the relatively facile and reproducible preparation of known stock eluent concentrations by direct weighing, compared to the more laborious preparation of C0,-free hydroxide eluents and necessary standardization. With efficient columns specifically developed for use with hydroxide eluents now available, if prepackaged COz-freehydroxide eluents in standard concentrations become commercially available, there would be little justification for continuing with any eluents other hydroxide for suppressed anion chromatography, whether gradient or isocratic.
ACKNOWLEDGMENT We thank Dionex Corp. for the use of their equipment and columns. Joan M. Slep (Celanese Corp.) is thanked for the generous gift of the Celgard hollow fibers of various types. LITERATURE CITED (1) Dasgupta, P. K. I n Ion Chromatograpby; Tarter, J. G., Ed.; Marcel Dekker: New York. 1986: DD 191-367. (2) Sunden, T.; Linaren, M.: Cederaren, A.; Siemer, D. D. Anal. Cbern. 1983, 55,2-4.(3) Sunden, T.; Cedergren, A.; Siemer, D. D. Anal. Cbem. 1984, 5 6 , 1085-1089. (4) Siemer, D. D.; Johnson, V. J. Anal. Chern. 1984, 5 6 , 1033-1034.
800
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(5) Tarter, J. G. Anal. Chem. 1984, 5 6 , 1264-1268. (6) Dasgupta, P. K. Anal. Chem. 1984, 56, 769-772. (7) Irgum. K. Anal. Chem. 1987, 59, 358-362. (8) Riviello, J. M., Pohl, C. A,, Taylor, M. S.37th Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy, Atlantic City, NJ, March 1986; Abstract No. 447. (9) Rocklin, R. D.; Pohl, C. A. 37th Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy, Atlantic City, NJ, March 1986; Abstract No. 585. (10) Rein,; M I Van der Linden. W E , Poppe, H Anal Chjm Acta 1981, 123 229-237 (11) Dasgupta. P. K. Anal. Chem. 1984, 5 6 , 96-103. (12) Stilliin, J. R. LC Mag. 1985. 3 , 802-812. (13) Mercurio-Cason, M. A.; Dasgupta, P. K.; Blakeley, D. W.; Johnson, R. L. J . Membr. Sci. 1986. 2 7 , 31-40. (14) Tanaka, K.; Ishihara, Y.; Sunahara, H. Bunseki Kagaku 1975, 2 4 , 235-238. (15) Girard. J. E. Anal. Chem. 1979, 57, 836-839.
(16) Tarter, J. G. J . Liq. Chromatogr. 1984, 7 , 1559-1566. (17) Ruricka, J.; Hansen, E. H.; Zagatto, E. A. Anal. Chim. Acta 1977, 88, 1-36. (18) Stumm, W.; Morgan, J. J. Aquatic Chemistry, 2nd ed.; Wiley: New York, 1981; p 212. (19) Dasgupta, P. K.; Bligh, R . Q.; Lee, J.; D'Agostino, V. Anal. Chem. 1985, 5 7 , 253-257. (20) Hwang, H.; Dasgupta, P. K. Anal. Chem. 1986, 5 8 , 1521-1524.
RECEIVED for review September 3,1986. Accepted November 24,1986. This research was supported by the Office of Basic Energy Sciences of the U.S. Department of Energy under Grant No. DE-FG05-84ER-13281. However, this report has not been subject to review by the agency and no official endorsement should be inferred.
Computer-Controlled Pneumatic Amplifier Pump for Supercritical Fluid Chromatography and Extractions Gilbert L. Pariente, Stephen L. Pentoney, Jr., Peter R. Griffiths,* and Kenneth H. Shafer
Department of Chemistry, University of California, Riverside, California 92521
A computer-cuntrdled pumping system for supercritkai fluids has been constructed based on a pneumatic amplifier pump. When CO, Is used as the fluM, It has been shown that the pump has pressure control as good as that of a syringe pump. I t must refill more often than a syringe pump with a large syringe volume, but the reflil t h e is short enough that the pump can refill during supercritical fluid extraction or chromatography without 111 effect. Additionally, the pump can attaln a set pressure much faster than syringe pumps, whlch makes it much more convenlent for extraction experiments where several dlscrete pressure intervals may be desirable.
The advantage of using supercritical fluid chromatography (SFC) for the high-resolution separation of molecules of low volatility has become widely recognized in recent years. The burgeoning popularity of SFC stems from the advantages that may be gained from the density, solvation, and diffusivity characteristics of supercritical fluids. A supercritical fluid can solvate large molecules that cannot be volatilized in a gas chromatograph (GC). The diffusivities of solutes in supercritical fluids are greater than their diffusivities in liquids. Thus the equilibrium of solutes between the mobile phase and the stationary phase is more rapid than in high-performance liquid chromatography (HPLC). In addition, the viscosity of most supercritical fluids is sufficiently low that they may be used with long open tubular columns to achieve higher resolution than HPLC using packed columns in reasonable analysis times. A further advantage of supercritical fluids over liquid solvents is that it is possible to control the solvation of solutes in the supercritical fluid by varying the density of the fluid. This control is accomplished by varying the pressure and/or the temperature of the system. This property makes supercritical fluid extraction (SFE) especially attractive in view of the potential selectivity of extraction by pressure control. Until recently no commercial instrumentation for capillary SFC has been available. Modified Varian 8500 syringe pumps (I,2) have been used by many workers for capillary SFC, and recently other syringe pumps specifically designed for SFC
are beginning to be introduced commercially ( 3 ) . Syringe pumps allow direct control of the flow rate of the mobile phase, while pressure is controlled by increasing or decreasing the flow rate through the restrictor at the end of the column. Refilling syringe pumps can be a time-consuming procedure. While it may not be necessary to refill a syringe pump often for capillary SFC because of the low flow rates required (a few microliters per minute), when supercritical fluids are used for packed column chromatography or for extractions where flow rates of several milliliters per minute may be desired, the slow speed of the refill cycle presents a severe limitation, since a reduction of pressure can occur for several minutes each time the pump refills. Reciprocating pumps also have been modified to pump compressible fluids above their critical pressure in an attempt to circumvent this limitation of syringe pumps ( 4 , 5 ) . Reciprocating pumps have enjoyed less popularity than syringe pumps in part because the pump heads must be cooled to improve pumping efficiency of fluids such as carbon dioxide ( 4 , 5 )and in part because of the large pressure fluctuations due to the reciprocation of the piston ( 5 ) . Both types of pumps are primarily designed for flow control and must be modified in order to control the pressure of the mobile phase. Conversely, pneumatic amplifier pumps are designed to contol output pressure and should therefore have appropriate properties for pumping compressible fluids. This type of pump works by applying air pressure to the lowpressure side of a two-sided pump piston. The low-pressure side of the piston has a large area relative to the side that displaces the fluid to be pumped (high-pressure side). To refill the pump, the pressure is removed from the low-pressure side and the piston is allowed to slide back while fluid from a reservoir flows into the empty chamber. Past applications of these pumps have been successful but cumbersome. These systems controlled output pressure by regulating the flow of high-pressure nitrogen into the low pressure side of the pump by means of a pressure regulator. Pressure programming was accomplished by attaching a stepper motor to the regulator (6, 7). In this paper we describe a new method of controlling a pneumatic amplifier pump for SFC and SFE. This pump
0003-2700/87/0359-0808$01.50/0 C 1987 American Chemical Society
