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Anal. Chem. 1988, 60, 1666-1669
Redox Suppressor for Ion-Exclusion Chromatography of Carboxylic Acids with Conductometric Detection Tetsuo Okada
Faculty of Liberal Arts, Shizuoka University, 836 Ohya, Shizuoka 422, Japan
A redox suppression scheme for Ion-exclusion chromatography of carboxylic acids wlth conductometrlc detectlon Is described. Hydrlodlc acld Is used as an eluent followed by postcolumn reaction with hydrogen peroxide. The redox suppressor (postcolumn reactor) was a 10 m X 0.2 mm 1.d. reactlon coil malntained at 70 O C wlth 1.1 M H202 belng mlxed wlth the column effluent at 0.4 mL/mln. The peak heights of some weak carboxylic adds obtained by d n g this suppressor were up to an order of magnRude higher than those obtalned wlthout the suppressor. Thls suppressor was partlcularly useful wlth h w y acldlc eluents, whlch permH a large number of carboxyllc aclds to be resolved by Ion-exciuslon chromatography. Thls suppression scheme can reduce background conductance more effectlvely than a conventlonal Ion-exchange suppressor. With a 4 mM strong monoprotlc acld as eluent, the Improvement can be more than a factor of 2 compared to catlon exchange wlth tetrabutylammonlum Ion.
High-performance liquid chromatography is a common and useful tool for determining organic acids. In order to separate organic acids, many separation modes, such as ion-pair chromatography on reversed phases (1-3), ion-exchange or ion-exclusion chromatography (IEC) with anion- or cationexchange resins (4-8), etc., have been investigated by many workers. IEC is particularly useful because it can easily distinguish organic acids from each other and from inorganic anions of strong acids (6-8). Ultraviolet absorptiometry, potentiometry, and conductometry have been investigated for use in IEC (9-15). Conductometric detection has become particularly sophisticated during the last decade with the development of ion chromatography (11-15) and is also particularly useful for measuring organic acids. Conductometric detection of organic acids following ion-exchange or ion-exclusion chromatography has been described by many researchers (7, 8, 12, 16). In ionexchange chromatography, with columns of typical exchange capacity currently in use, a relatively high ionic strength eluent is required in order to elute the analyte ions within an acceptable time period. In IEC, relatively strong acid eluents are required to simultaneously isolate many organic acids. However, these elution conditions are not ideally compatible with conductometric detection because low background conductance is necessary for sensitive detection. Attempts to reduce the background conductance for conductometric detection of organic acids in IEC have thus far involved precipitation and ion-exchange suppression schemes. In the first, a halogen acid, e.g., HC1, is used as eluent and an Ag+-form cation-exchange column is used as the suppressor. The suppression reaction involves the replacement of H+ by Ag' and consequent formation and precipitation of AgCl ( 17). Unfortunately, precipitation in the suppressor column causes increased back pressure. Although periodically cutting off the spent portion of the plastic suppressor column ameliorates the problem to some degree, it is not a convenient solution. The second scheme involves simply ion-exchanging the H+ for another cation, e.g., Na+ or K+, and thereby reducing the
background conductance (8). Further developments along this line have led to the use of octanesulfonic acid, tridecafluoroheptanoic acid, or perfluorobutyric acid as eluents followed by postcolumn exchange of H+ for NBu4+by using a cation-exchange membrane-based suppressor (18,19). While the technique thus permits continuously regenerated membrane-based suppression, with a background conductivity typically ranging from 45 to 55 N/cm for 1mM octanesulfonic acid (20), there is room for improvement, particularly if eluents of higher acidity must be used. In IEC, the eluent pH should be lower than the pK, values of the analyte acids of interest to obtain good chromatographic resolution. The pK, values of most carboxylic acids fall between 2 and 5. Thus, the eluent pH should be lower than 3; that is, for a strong acid eluent, the eluent concentration should be higher than 1 mM. In this paper, I explore a redox suppression scheme for IEC of carboxylic acids. EXPERIMENTAL SECTION The chromatographic system was composed of the following components: two computer-controlled pumps (CCPD), a column oven (CO-SOOO), a conductometric detector (CM-8000) (all from Tosoh Co.,LM.), an injection valve equipped with a 100-pLsample loop, a static mixer (sequentially dockwise- and anticlockwisewound poly(tetrafluoroethy1ene) (PTFE) tube coil, 0.2 mm i.d., 12 m in length), and a reaction coil (PTFE tube, 0.2 mm i.d., 5-15 m in length). The separation column was TSKgel SCX (7.6 mm i.d. X 30 cm, H+-form cation-exchange resin). The column and the detector were kept at 35 "C in the oven. Effluent pH was monitored by a pH meter (Model HMBS, Toa Electronics Co., Ltd.) equipped with a glass electrode, calibrated by the two-point method. Standard solutions of organic acids were prepared by dissolving analytical grade organic acids in distilled deionized water. A hydriodic acid eluent was prepared fresh daily by diluting analytical reagent grade hydriodic acid in distilled deionized water and deaerating the solution. Hydrogen peroxide solutions were prepared by diluting analytical grade HzOzin distilled deionized water and were standardized by titration with secondary standard potassium permanganate (standardized with oxalate).
-
RESULTS AND DISCUSSION A Redox Reaction for Background Suppression. The following reaction was used for the present redox suppression scheme: 21-
+ 2H+ + HzOz
-
I,
+ 2H20
Hydriodic acid was used as the eluent and hydrogen peroxide as the postcolumn reagent. If the suppressor reaction occurs adequately, the background conductance should be significantly reduced. A substantial decrease in conductance was indeed observed in preliminary studies. Further, an excess of HzOz,a very weakly ionized substance, did not significantly affect the background conductance; in fact it promoted the extent of the suppressor reaction. Therefore, the conditions for this reaction were optimized as follows. Optimization of the Suppressor Reaction. The reaction conditions to be optimized were reaction temperature, the reaction time (represented by the length of a reaction coil at a fixed flow rate), and the concentration and flow rate of the
0003-2700/88/0360-1666$01.50/0 0 1988 American Chemical Society
ANALYTICAL CHEMISTRY, VOL. 60, NO. 17, SEPTEMBER 1, 1988
Table I. Effects of Reaction Temperature and Length of Reaction Coil on Background Conductancea temperature, "C
conductance, rS/cm
30 40 50 60 70 80
517 398 254 131 113 118
Postcolumn reaction with 1.1 M HzOz flowing at 0.4 mL/min; 10-mreaction coil. postcolumn reagent, H2OP The optimum flow rate of the eluent (14mM) was previously determined to be 0.8 mL/min for the chromatographic separation. Reaction Temperature. The variation of the background conductance with the reaction temperature (30-80 " C ) is shown in Table I. The conductance of a 4 mM HI solution is 1440 pS/cm. The conductance of the reactor effluent decreased with increasing reaction temperature and reached 114 pS/cm at 70 "C. A slight increase in the conductance was observed by increasing the temperature to 80 "C, likely due to an increase in ionic mobility. Thus, the data in Table I shows the suppressor reaction is completed to the maximum attainable extent by 70 "C. The base-line noise decreased with increasing reaction temperature up to 70 "C. At low temperatures, the high base-line noise resulted from the high background conductance itself as well as from the variability of the largely incomplete extent of the suppression reaction. The noise observed at 70 "C was one-twentieth the magnitude of that at 30 "C. Traces of ions such as molybdate are known to catalyze the H202-Ireaction (21);however, no significant benefits were realized by incorporating traces of molybdate in the Hz02reagent in the low-temperature studies. Reaction Time. The effect of reaction time was studied by varying the length of the reaction coil. The use of a 10-m reaction coil permitted the background conductance to be reduced to 114 pS/cm at 70 "C; the background conductance obtained with 5-m and 15-m reaction coils was 140 &/cm and 115 pS/cm, respectively, at the same temperature. The eluent reacted at room temperature with the reaction reagent for about 18 s in the mixer followed by another -15 s during passage through the 10-m reaction coil. The suppressor reaction was not completed with a 5-m coil, while a 10-m coil was sufficient to reduce the background conductance of a hydriodic acid eluent at a reaction temperature of 70 "C. Concentration and Flow Rate of the Reaction Reagent. The concentration and the flow rate of the H202solution were optimized with a 10-m reaction coil maintained at 70 "C. With 0.28 M H202at 0.8 mL/min, the background conductance decreased from 740 pS/cm for unsuppressed 2 mM HI to 140 pS/cm. The background conductance decreased further as the concentration and the flow rate of the reaction reagent were increased and became essentially constant with a H202 concentration of >0.55 M at a flow rate of >0.4 mL/min. Table I1 shows the dependence of the background conductance and pH as a function of H202concentration and flow rate. The conductance of the effluent from the redox suppressor decreased with an increase in either the flow rate or the concentration of H202. However, the effluent pH could not be increased beyond 3.6 for the 4 mM HI eluent and 3.77 for the 2 mM HI eluent. The formation of triiodide from iodide and molecular iodine (I2 I- = Is-) is well-known. This reaction must occur in the present suppressor system and prevents complete conversion of iodide to iodine in the redox suppressor. An anionic species (Is-)remains together with hydrogen ion after the reaction.
+
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Table 11. Effects of the Concentration and Flow Rate of Reaction Reagent on the Effluent pH concn of eluent, mM
concn of
Hz02, M 0 0.28
4
1.1
2.2 0 0.28 1.1
2
2.2
flow rate, mL/min
conductance, pS/cm
pH
0 0.8 0.2 0.8 0.2 0.8 0 0.8 0.8 0.2 0.8
1440 285 574 101 134 96 740 140 79 97 75
2.45 3.16 2.87 3.63 3.61 3.62 2.72 3.49 3.77 3.71 3.76
I
u 3
OO
0.L
Flow Rate of
HzO2, m l l n n
Figure 1. Variation of peak heights of tartaric and acetic acids with the flow rate of reaction reagent. Broken lines show the decreases in the peak heights calculated by taking into account the dilution. Conditions: sample concentration, 50 ppm; eluent, 2 mM HI; reagent,
1.1 M H,02.
Hydrogen ion is the only cation that exists in the present system. Therefore, the above reaction interferes with the completion of the suppressor reaction. However, this reaction promotes the solubility of iodine. Iodine resulting from the suppressor reaction of the 4 mM HI eluent may have led to precipitation because of the poor solubility of iodine (1.5 X low3M at 30 " C ) if the above reaction did not occur. The calculated pH of 4 mM HI is 2.43, based on -yH+ calculated according to the Debye-Huckel equation for this ionic strength. The measured pH of this eluent was 2.45 (Table 11). For 4 mM HI (0.8 mL/min) reacting with 1.1M H202(0.2 mL/min), the effluent pH is 3.61. Again assuming the applicability of the Debye-Huckel equation (dilute solutions of uni-univalent electrolytes), the effluent [H'] is computed to be 2.49 X M. Once the volumetric dilution is accounted for, it may be calculated that 92.2% of the H+ (or HI) has been removed. Similarly for the 2 mM HI eluent, after activity corrections, the calculated pH of 2.72 exactly agrees with the measured pH. After reaction of the eluent (0.8 mL/min) with 1.1M H202at 0.2 mL/min, [H+] is calculated from the measured pH after activity corrections to be 1.98 X lo-' M. This corresponds to 87.6% of the HI being removed. For further improvement of the redox suppressor, some means of removing iodine resulting from the suppressor reaction may be necessary. This remains to be investigated in the future. Figure 1shows the variation of the peak heights of tartaric and acetic acids with the concentration and the flow rate of the Hz02solution. The decreases in the peak heights with increasing flow rate of the reagent were apparently caused by
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ANALYTICAL CHEMISTRY, VOL. 60, NO. 17, SEPTEMBER 1, 1988
Table 111. Comparison of Attainable Conductance Obtained by the Present Redox Suppressor with a Cation-Exchange Suppressor conductance, rc.S/cm eluent, mM
without suppressor, measd"
with redox suppressor, measd
4 2 1
1440 740 380
114 86 60
concn of
with ion-exchange suppressor, calcdb Na+ exchange NBu,+ exchange 384 192 96
508 254 127
a From infinite dilution mobilities, the conductance values for 1, 2, and 4 mM HI are calculated to be 427,853, and 1706, respectively. *If dilution of the column effluent occurred in this system to the same extent it does in the redox suppressor, these values would be reduced by the factor 0.67.
€
-0a 2+ €5
1stE
'
E
ol
r 1
a '
a
1o
c
L d 0 1 conc of H,Oz,
~
A 2 M
Flgure 2. Variation of peak height of tartaric acid with the concentration of hydrogen peroxide solution. Flow rate of the reaction reagent: (1) 0.2 mL/min, (2) 0.4 mL/min, and (3) 0.8 mL/min.
the dilution resulting from the reagent addition. The broken lines in Figure 1represent the calculated decreases in the peak heights solely due to the dilution. The peaks obtained at higher flow rates of H202 were larger than those for the calculated values because these conditions resulted in an increase in the effluent pH and thus increased the dissociation of these acids. I t may seem that these results suggest that a large excess of Hz02is beneficial. However, a large excess of H202actually decreases sample response; Figure 2 shows how the peak height of tartaric acid decreases with increasing concentration of H202 at three different reagent flow rates. Since this decrease cannot be attributed to dilution and the background conductance, if anything, decreases with excess Hz02,it is possible that excess H202causes some oxidative decomposition of the sample analytes. Moreover, high concentrations of H202 generated oxygen gas bubbles in the detector and thus caused base-line instability. Consequently, it was desirable to use 1.1 M H202at 0.4 mL/min as the reagent for the redox suppressor. Effectiveness of the Redox Suppressor in Detecting Carboxylic Acids. Table I11 shows the comparison of the background conductance values obtained by the present redox suppressor with those calculated from limiting ion equivalent conductance data for a cation-exchange suppressor if either Na+ or NBu4+ is exchanged for H+. Admittedly, the ionic strength of these effluents is too high for the use of infinite dilution equivalent conductances to be accurate. Nevertheless, the background conductance obtained with an ion-exchange suppressor will increase more or less linearly with the concentration of the eluent. As the eluent concentration increases, the advantage of the redox suppressor becomes significant. For the 4 mM HI eluent, the redox suppressor decreases the background conductance at least 2-fold better than if ion exchange for NBu4+were applied, even after accounting for
Figure 3. Improvement of resolution of organic acids by using highly acidic eluent. Peak identification: male, maleic acid; malo, malonic acid: cit, citric acid; gly, glycolic acid; for, formic acid; fum, fumaric acid. Sample concentration was 50 ppm.
dilution. Thus, the redox suppressor is particularly useful when highly acidic, low-pH eluents are needed. Concerns always exist in postcolumn reaction systems regarding band broadening. Using similar tubular reactors of even larger holdup volume, Vrdtny et al. reported that the main source of band broadening was the separation column, where 6 0 4 5 % of the total peak variance occurred, and that the contribution of the postcolumn reaction system was 1520% of the total variance (22). In this study, marked peak broadening was not observed and the band broadening caused by the redox suppressor was not a problem. Figure 3 shows the effectiveness of using highly acidic eluents in IEC. Malonic acid (pK, = 2.82; pK2 = 5.70) and citric acid (pK1 = 3.13; pK2 = 4.76, pK3 = 6.40) were not separated with 1 mM HI eluent (pH 3) but could be resolved by using 4 mM HI eluent (pH 2.45). The separation of glycolic acid (pK = 3.83), from formic acid (pK = 3.75) and fumaric acid (pK, = 3.10; pK, = 4.60) was also improved by using the more acidic eluent. These results show the utility of highly acidic eluents for the IEC separation of a multitude of carboxylic acids, and the present redox suppressor makes the conductometric detection possible with such eluents. Figure 4 shows chromatograms of nine carboxylic acids obtained with and without the redox suppressor. Peaks of all the sample acids increased with the use of the redox suppressor, regardless of the eluent concentration. The improvement in sensitivity is most notable with the weaker acids such as acetic acid (pK = 4.761, propionic acid (pK = 4.87), n-butyric acid (pK = 4-81),and isobutyric acid (pK = 4.84).
ANALYTICAL CHEMISTRY, VOL. 60, NO. 17, SEPTEMBER 1, 1988
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In conclusion, this redox suppression scheme effectively reduces the background conductance of a highly acidic eluent, which permits one to resolve many organic acids by IEC and also allows sensitive conductometric detection of organic acids.
ACKNOWLEDGMENT I wish to thank Toyo Soda Manufacturing Co., Ltd., for providing the separation columns. LITERATURE CITED (1) (2) (3) (4) (5) (6) (7) (8) (9) (10) (11)
Figure 4. Effectiveness of the redox suppressor system. c6omatograms were obtained without redox suppressor; lower chromatograms were obtained with the redox suppressor. Eluent concentration: (A) 1 mM; (B) 4 mM. Peak identification: (1) oxalic, (2) tartaric, (3) malic, (4) glycolic, (5) formic, (6) acetic, (7) propionic, (8) isobutyric, and (9) n-butyric acids. Sample concentration was 50 ppm except for iso- and n-butyric acids (200 ppm).
(12) (13) (14) (15) (16) (17) (18)
The peak heights of these acids obtained with the redox suppressor were an order of magnitude larger than those obtained without the suppressor for 4 mM HI as eluent. Further, the redox suppressor significantly reduced the water dip caused by the sample injection. As can be seen in Figure 4, oxalic acid, which eluted together with the injection dip, was detectable with the redox suppressor because of the reduction of the injection dip.
(19) (20) (21) (22)
Cassidy, R. M.;Elchuk, S. Anal. Chem. 1985, 57, 615-620. Miwa, H. J. Chromatogr. 1985, 333, 215-219. Skeily, N. E. Anal. Chem. 1982, 54, 712-715. Buytenhuys, F. A. J. Chromatogr. 1981, 218, 57-64. Rokushika, S.; Sun, 2. L.; Hatano, H. J. Chromatogr. 1982, 253, 87-94. Giod, E. K.; Kemuia, W. J . Chromatogr. 1988, 366, 39-50. Itoh, H.; Shinbori, Y. Chem. Lett. 1982, 2001-2002. Singsby, R. J. Chromatogr. 1986, 377, 373-382. Haddad, P. R.; Alexander, P. W.; Trojanowicz, M. J. Chromatogr. 1984, 315, 261-270. Egashira, S. J. Chromatogr. 1980, 202, 37-43. Small, H.; Stevens, T. S.; Bauman, W. C. Anal. Chem. 1975, 47, 1801-1 809. Okada, T.; Kuwamoto, T. Anal. Chem. 1983, 55, 1001-1004. Okada, T.; Kuwamoto, T. Anal. Chem. 1985, 57, 258-262. Okada. T.; Kuwamoto, T. Anal. Chem. 1985, 57, 829-833. Okada, T.; Kuwamoto, T. Fresenius' 2. Anal. Chem. 1986, 325, 883-685. Thurman, E. M. J. Chromatogr. 1979, 785, 625-634. Rich, W.; Smith, F.; McNeil, L.; Sidebottom, T. I n Ion Chromatographic Analysis of Environmental Pollutants; Muiik, J. D.,Sawiciki, E.. Eds.; Ann Arbor Science: Ann Arbor, MI, 1979; Voi. 2, pp 19-29. Smith, R. E. Ion Chromatograohy Apolications; CRC Press: Boca Ra.. ton, FL, 1988; pp 64-71. Gjerde, D. T.; Fritz, J. S. Ion Chromatography, 2nd ed.; Huthig: New York, 1987; pp 235-251. Dionex Corp., personal communlcation, 1988. Vogei, A. I.A Textbook of Quantitative Inorganic Analysis, 3rd ed.; Longmans Green: London, 1961; p 363. Vrltny, P.; Brinkman, U. A. Th.; Frei, R. W. Anal. Chem. 1985, 57, 224-229.
RECEIVED for review July 13, 1987. Accepted April 15, 1988.
