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Anal. Chem. 1082, 54, 1558-1563
minescence-chromatographic technique. CONCLUSIONS The HPLC/MSRTP system development appears to have far-reaching implications for fundamental spectroscopic studies of luminescence, for the study of the nature of the micellar aggregate, and for improved selectivity in chromatographic analysis methods. It is a versatile and reliable means of routinely observing phosphorescence in fluid solution at room temperature, thus facilitating extensive photophysical studies of luminescent molecules. Because of the ease of adding concomitants with the chromatographic pumping system, it permits numerous luminescence probe studies of the effect of bulk solution conditions on the integrity of the micellar assembly. Competitive sensitivity is possible in analysis by utilizing total luminescence detection in some cases, and improved selectivity is possible by observation of the different relative phosphorescence enhancemenb. In other studies in our laboratory, the mere presence or absence of a phosphorescencesignal has been used to eliminate numerous possible solutes in complex coal samples for peak identification purposes. For the two modes studied, micellar chromatography and postcolumn reaction, each have their own unique features that can be applied to the needs of a particular analysis. For optimal sensitivity, the micellar chromatographic mode can be employed. For further enhancement of selectivity, the more chromatographically efficient postcolumn reaction mode is suggested if the resulting lower sensitivity can be tolerated. Thus, HPLC/MSRTP offers an adjustable device to increase the selectivity of analyses for compounds exhibiting the phenomenon. The list of substances known to phosphoresce a t room temperature is continually growing and practical applications for compounds which are inherently weakly fluorescent but strongly phosphorescent are expected soon.
ACKNOWLEDGMENT The authors thank Technicon, Inc., for the loan of some of the chromatographic equipment, Supelco, Inc., for aid in obtaining some of the columns, and Spectra Physics Co. for the loan of the fluorescence detector. LITERATURE CITED Fendler, J. H.; Fendier, E. J. ”Catalysis In Micellar and Macromolecular Systems”; Academic Press: New York. 1975. Tanford, C. “The Hydrophobic Effect”, 2nd ed.; Wiiey: New York, 1980. Thomas. J. K. Chem. Rev. 1980. 80. 283-299. Turro, N. J.; Gratzel, M.; Braun, A. M.’Angew. Chem., Inf. Ed. Engl. 1980, 19. 675-696. Cline Love, L. J.; Skrilec, M.; Habarta, J. G. Anal. Chem. 1980, 52, 754-759. Cllne Love, L. J.; Habarta, J. G.; Skrllec, M. Anal. Chem. 1981, 53. 437-444. Skrllec, M.; Cllne Love, L. J. Anal. Chem. 1980. 52, 1559-1564. Cline Love, L. J.; Skrilec, M. J. fhys. Chem. 1981, 85, 2047-2050. Armstrong, D. W.; Henry, S. J. J. Llq. Chromafogr. 1980, 3 , 657-662. Armstrong, D. W.; Nome, F. Anal. Chem. 1981, 53, 1662-1666. Snyder, L. R.; Klrkland, J. J. “Introductlon to Modern Llquid Chromatography”, 2nd ed.; Wlley: New York, 1979; Chapters 4 and 6. Snyder, L. R.; Klrkland, J. J. “Introduction to Modern Liquld Chromatography”, 2nd ed.; Wiley: New York, 1979; pp 740-746. Shaeiwltz, J. A.; Chen, A. F.-C.; Cussier, E. L.; Evans, D. F. J. Colloid Interface Scl. 1981, 8 4 , 47-56. Llanos, P.; Zana, R. Chem. fhys. Lett. 1980, 76, 62-67. Manabe, M.; Koda, M. Bull. Chem. SOC.Jpn. 1978, 57, 1599-1601. Yarmchuk, P.; Welnberger, R.; Cllne Love, L. J., submitted for publlcation in Anal. Chem,
RECEIVED for review March 8,1982. Accepted May 4,1982. This work was supported in part by the National Institutes of Health, Grant No. GM-27350. This work was presented in part at the 33rd Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy, Atlantic City, NJ, March 8, 1982. Abstract No. 83.
Dynamically Coated Columns for the Separation of Metal Ions and Anions by Ion Chromatography R. M. Cassldy” and Steven Elchuk General Chemistry Branch, Atomic Energy of Canada Limited, Research Company, Chalk River Nuclear Laboratories, Chalk River, Ontario, Canada KOJ IJO
A number of hydrophobic molecules contalnlng ionic functlonai groups have been dynamically coated onto 5 and 10 pm reversed phases, and these phases have been evaluated for the ion-exchange separatlon of cations and anions. Several sulfate (C12,C20)and sulfonate (C6, C,) cation systems and quaternary ammonium salt (c89 Cl6, C21, C32, C37) anion systems were examined and compared with respect to column efficiency. The potential of this approach for the preparation of ion exchangers of variable capacity is discussed and some associated analytical aspects are examined brlefiy.
Ion chromatography,f i s t described in 1975 (I),has become a popular technique for the determination of anions, usually a difficult analysis by classical techniques. The unique feature 0003-2700/82/0354-1558$01.25/0
of this system is the use of a suppressor column to convert the eluent ions into species of low conductivity and consequently the eluite can be detected very sensitively with a conductivity monitor. The suppressor column does impose some limitations, however, such as degradation of column resolution, the requirement for periodic regeneration of exhausted suppressor columns, variations in the retention time of some peaks (particularly water) with suppressor exhaustion, and limited choice of eluents. A further limitation is the fact that ion chromatography is not a high-performance system when compared to current state-of-the-art liquid chromatography. Consequently other chromatographic approaches have been investigatedfor the determination of anions (2-11). One approach is the use of low capacity ion-exchange materials (2-7) with small concentrations of eluents such as phthalate, benzoate, or citrate in the salt and/or free acid forms. Two 0 1982 American Chemical Soclety
ANALYTICAL CHEMISTRY, VOL. 54, NO. 9, AUGUST 1982
different column materials have been used for these studies; a macroporous styrenedivinylbenzene resin (2,5)and a surface quaternized silica (6, 7). The first material has been applied to the separations of both cations and anions but is not commercially available as a highperformance packing. The silica exchanger is available commercially and a recent study (8) has compared the resolution of this exchanger with that of the agglomerated latex e:ochangers used for classical ion chromatography (9). This silica exchanger is available as a 10-pm high-performance packing but is not available in cation form and cannot be used in high or low pH conditions. Another study (10) has shown that a weak cation exchanger bonded to silica can be used with eluent suppression to separate alkali cations. An alternate approach tal the analysis of inorganic ions has been the use of reversed-phase columns and ion pairing reagents such as cetyltrimethylammonium bromide, tetrabutylammonium hydroxide (10,11), and heptyl sulfate (10). Because reversed-phase columns are readily available and used by most high-performance liquid chromatography (HPLC) practioners, this approach is attractive. However, further studies are required to more completely evaluate this technique. The effects of variations in the molecular size of the modifier on column perforrnance is of interest since it should be possible to dynamically coat reversed phases with high molecular weight modifiers !whichwould not desorb in aqueous eluents. Such systems would not require the addition of the modifier to the mobile phase and should function as ion exchangers. Minor changes5 in coating procedures (solvent composition and/or modifier concentration) may permit the preparation of columns having a wide range of ion-exchange capacity. These systems chould be applicable to ion chromatography and ion separations with low conductivity eluents and to trace enrichment where the adjustment of the capacities of the column and enrichment cartridge can be important. This paper describes the results of a study of the behavior of styrenedivinylbenzene and bonded-C18high-efficiency reversed-phase columns coated with a variety of anionic and cationic modifiers. EXPERIMENTAL SECTION Apparatus. The HPLC Bystem consisted of a conventional HPLC gradient pumping system (M6000A, Waters Associates, Milford, MA), a Rheodyne injection valve (Model7120, Rheodyne, Berekeley, CA), a variable wavelength absorbance detector (Tracor, Austin, TX), and a conductivity detector (Laboratory Data Control, Riviera Beach, FL). The detector output was monitored with a data processing and disk storage unit (Model 81I.OBRl Bascom Turner Instruments, Newton, MA), and the chromatograms were processed with a computing integrator (SP41010, Spectra Physics, Santa Clara, CA). The values of height-equivalent to a theoretical plate (HETP) were calculated by the area method (12);although this is not the most accurate method for tho calculation of HETP values, it does give better results than tangent or peak height techniques, especially for skewed peaks (12). The eluted metal ions were detected after a postcolumn reaction with 4-(2-pyridylazo)resorcinol(PAR). The postcolumn reactant was added to the eluate with a special low-volumemixer (13)at the same flow rate used for elution of the metal ions, and the resulting solution was monitorled by absorption spectrophotometry at 530-540 nm. Reagents and Materials. The PAR solutions (2 X mol-L-') were made 2 mol.L-Lin ammonia and 1 mo1.L-1 in ammonium acetate. For metal ion separations the eluent WBS purified by a combinationof ion exchange and constantawrent electrolysis at 2.5 mA for 24 h over a Hg-pool electrode. All aqueous solutions were prepared with distilled water purified with a Milli-Q system (Millipore Corp., Bedford, MA). The ion-pairing reagents used in this study are listed in Table I. The sodium salts of the siilfonatesand sulfates were obtained from Regis Chemical Co. (Morton Grove, IL), and the three
1559
Table I. Ion-Pairing Reagents name
structure
(a) Cation Exchange Reagents 1-hexanesulfonate CH3(CH2)SS031-octanesulfonate CH,(CH, ),so,1-dodecylsulfate CH3(CH2)1Is,'1-eicosylsulfate CH3(CHZ)l (b) Anion Exchange Reagents tetraethylammonium (CH,CH,),N+ salt tetrabutylammonium (CH,(CH,),),N+ salt trioctylmethylCH,(CH,(CHJ,),N+ ammonium salt tetraoctylammonium (CH,(CH, ), ),N+ salt tridodecylmethylCH,(CH,(CH,),,),N+ ammonium salt
no. of carbon atoms ' 6
C8 Cl, C20
c* 6I'
c2, c32
C,,
a The anion reagents were used as the chloride, iodide, or hydroxide form.
lower-molecular-weightquaternary salts were obtained from Aldrich Chemical Co. (Milwaukee,WI). The tetraoctylammonium iodide was provided by P. Leeson, Chalk River Nuclear Laboratories, and the tridodecylmethylammoniumiodide was prepared by refluxing 40 g of tridodecylamine with 25 g of methyl iodide in 200 mL of 1:l acetonitrile-methanol for 1.5 h. The product separated as an oil when water was added and a white solid salt was obtained by crystallization (three times) from diethyl ether solutions placed in a freezer at 253 K. The reversed phases used were 5-pm C18(Supelosil,Supelco, Bellefonte, PA) and 10-pm C18phases bonded to silica (Partisil PXS 10125ODs-3, Whatman, Clifton, NJ; and MCH-10, Varian Associatea, Walnut Creek, CA), and a 10-pmstyrenedivinylbnzene phase (PRP-1, Alltech Associates, Deerfield, IL). Column Coating. Large hydrophobicmodifiers were dissolved in methanol-water or acetonitrile-water mixtures to give to mo1.L-l solutions, and these solutions were passed through the column until equilibration was achieved, at which point the mobile phase was switched to water. The volume required for equilibration varied with the solvent composition and the concentration of the modifier. The parameters used for the examples discussed in this report are given in Results and Discussion. RESULTS AND DISCUSSION Cation Exchange Systems. Uniformly coated Czosulfate columns were prepared by the passage of 500 mL of a 2.5 X 10" mo1.L-l solution in 25% (v/v) acetronitrile-75% water which was then followed by deionized water; further coatings produced only minor changes in retention times. These columns were normally used with aqueous mobile phases that did not contain any of the Czosulfate because the sulfate was adsorbed strongly by the bonded phase; even small concentrations (lo4 to 10" mol-L-l) in the mobile phase were absorbed onto the column and this buildup eventually caused significant increases in the pressure drop across the column. An example of the rapid metal ion separations that can be obtained with these C20sulfate columns is shown in Figure 1. The peak symmetry was generally better than that found previously by the authors for bonded-phase ion exchanges and styrenedivinylbenzene ion exchangers (11,13,14). Column efficiency was studied for three concentrations of the tartrate eluent (0.045, 0.06, and 0.075 molmL-'), and data for one of these concentrations are shown in Figure 2. Only minor changes in column efficiencies were observed for the different tartrate concentrations, and these may be due to mass transfer effects arising from variations in the distribution of the different metal species with changes in the concentration of the tartrate ligand. A direct comparison of these column effi-
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ANALYTICAL CHEMISTRY, VOL. 54, NO. 9, AUGUST 1982
F
t
Pb
0.10~ cu
M"
1
I
E E
p
0.05
Zn
Li
I
co
L 0
0
50
RETENTION TIME Is1
Flgure 1. Separation of metal ions on a 10-pm C1, column previously equilibrated with C,,H,,SO,Na. Experimental condltlons: Varian column, 30 cm; eluent, 0.075 mo1.L-l tartrate (pH 3.4) at 2 mL-min-'; sample, 20 pL of 5 ppmL-l solutions of Mn(II), Co(II), and Cu(I1); detection, absorption at 530 nm after postcolumn reaction. 0.3
-e E
Mn CO
cu "I!-
LINEAR VELOCITY (mm s
-' I
Flgure 2. Column efficiency study for 10-pm C1, column previously equilibrated w h C2,H,,SO,Na. Experimental conditions: eluent, 0.045 mo1.L-l tartrate (pH 3.4); other conditions, as for Figure 1.
ciencies with other data reported for metal ions is difficult because column efficiency for any one column can vary with the nature of the metal ion, the nature of the eluent (usually a complexing reagent), and the concentration of the eluent species. However it does appear that this 10-pm system gives HETP values that are more consistent from metal ion to metal ion and HETP curves that have a smaller slope than other HPLC ion exchange systems studied by us (12,14,15). The metal ions appeared to be separated by an ion exchange mechanism on columns coated with this large Czosulfate rather than by ion pairing, because the mobile phase did not contain any of the CzOsulfate. The addition of small concentrations of the sulfate to the mobile phase had no noticeable effect on the separation; continued use led to band broadening which was attributed to a build up of the sulfate on the top of the column. After continuous use of a CzOsulfate column for more than 1week no changes were observed in retention times of column efficiency. The retention of the sorbed Czosulfate over a longer term was not studied since the sulfate could be quickly stripped with 70% (v/v) acetonitrile-30% water and then recoated. The longer term stability of the C18bonded phases is important, however, and it was found that the bonded phase was stable to continuous use for 1month; longer term stability was not investigated. Because other reports have shown that small amounts of a polar organic solvent added to an aqueous mobile phase can increase the available surface area for hydrophobic supports (16),the effect of small amounts (up to
I
2
LINEAR VELOCITY (rnrn.s
3
4
-' 1
Flgure 3. Column efficlency study for 5-pm C18column in equilibrium with C6H,,S0,Na. Experimental conditions: Supelcosil column, 15 cm; eluent, 0.045 mo1.L-i tartrate (pH 3.4) and 0.01 mo1.L-l C&i&O~Na; detection, as for Flgure 1.
5% of v/v) of methanol on this hydrophobic system was studied; the results showed that these additions had no appreciable effect on column efficiency or retention times. Columns coated with the C12sulfate gave metal ion separations similar to those obtained with the Czosulfate system in terms of column efficiency and selectivity; however, the hydrophobic portion of the C12sulfate was not large enough to prevent losses from the column into the eluent and small concentrations of the sulfate had to be added to the mobile phase. For a 30-cm C18 bonded phase in equilibrium with a lo4 mo1.L-' aqueous solution of the C12sulfate, approximately 240 mg of the sulfate was sorbed, and the column required 0.15 mo1.L-' tartrate eluent (pH 3.4) for elution of the metal ions with retention times similar to those shown in Figure 1. Aqueous solutions of the C6 and C8 sulfonates equilibrated rapidly with the C18column and these systems gave separations similar to those obtained with Clz and CZ0sulfates. With a mo1.L-l solution of the C8 sulfonate, equilibration was reached after passing 125 mL through the column. Good separations were obtained over the ramge of 10-1 to mo1.L-l for the C6 sulfonate, but at mo1.L-l there was a rapid deterioration in column efficiency possibly due to low column capacity or poor wetting of the hydrophobic support. The C8 sulfonate could be used at concentrations of mol-L-' due to its stronger sorption onto the support. The column efficiencies observed with these modifiers were better than the longer chain sulfates by 50 to 90%; HETP values were generally in the range of 0.08-0.15 mm at 1 ml-rnin-l. Whether this improved efficiency is related to the chain length, the difference in the functional groups, or difference in the uniformity of the sorbed modifier is unclear at present. Figure 3 shows the results from a column efficiency study of a 5-p m bonded phase in equilibrium with C6HI3SO3Na. As expected, the performance of this column is superior to that observed for the 10-pm packing (Figure 2) and the excellent performance of this system is further illustrated by the isocratic separation of six metal ions shown in Figure 4. The results shown in Figures 3 and 4 illustrate that metalligand equilibria do not limit the attainment of the rapid mass transfer required for high-performance separations. However, some observations did suggest that metal-ligand equilibria could be important. The latter part of the Mn(I1) peak in Figure 4 has a small shoulder which appeared as a second peak a t low flow rates. The ratio of the two peaks changed with flow rate and with the miount of Mn(I1) injected; several testa indicated that impurities and/or sorption onto silanol groups were not responsible for this behavior. This phenomenon,
ANALYTICAL CHEMISTRY, VOL. 54, NO. 9, AUGUST 1982
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r
I
1
0'3
I
I
I
I 2
4
3
LINEAR V E L O g u m m . r - ' I
RETENTION TIME lrnlnl
Figure 4. Separation of six irietal ions on a 5-pm Cia column In equilibrium with CBHi,S03Na. Experimental conditions: Supelcosil column, 15 cm; eluent, 0.045 rnio1.L-l tartrate (pH 3.4)and 0.01 mo1.L-l C,Hi3S0,Na at 1.5 ml-min-'; sample, 20 pL of a solution containing 25-50 pg-mL-l of each metal ion; detection, as for Figure 1. which may be due to slow equilibrium between different Mn(I1) complexes, was not examined further since it was not of interest to the main purpose of this study. The capacities of the C1:! and (220 column systems studied varied from about 0.1 to 1nnequiv for 30-cm column (column volume 3.8 mL) as determined by the weights of absorbed modifier obtained when the columns were stripped with 70% (v/v) acetonitrile-30% water. A conventional styrenedivinylbenzene ion-exchange! resin would be expected to have a capacity of -4-6 mequiv for an equivalent column volume. Commercially avails ble boinded-phase cation exchangers are often listed as having Capacities in the 1 mequivg-l range; consequently they should give columns of the same capacity as conventional resins because these silica based exchangers have larger densities. For two exchangers having chemically similar exchange sites, the eluent concentration required for elution of a series of eluiteo in a similar time span should be approximately the same for equal exchange capacities; however, in the authors' experience, smaller concentrations of eluents (by a factor of 10) were usually required for elution of metal ions from silica based exchangers. Consequently it would appear the effective capacities of silica based exchangers are much smaller than 1mequivng-', and likely in the same range as the dynamically prepared exchangers studied here. The capacity of the dynamically prepared exchangers can be varied, and this is an attractive feature for many applications. Anion Exchange Sysitems+l8 Bonded Phases. A number of quaternary ammonium salts ((28, C16, C25,C32, Cg7 compounds shown in Table I) were used to prepare dynamically coated columns for the separation of anions. For the higher molecular weight quaternary salts (C%,C32, CS7)coated columns were prepared by the establishment of equibrium in a methanol-water solvent. The amounts coated on the columns, which varied from 20 to 80 mg, depended on the quaternary salts, the colurnin, and the solvent composition; for a 6.67 X lod4 mo1.L-' solution of tridodecylmethylammonium iodide in 80% (v/v) methanol-20% water, the equilibrium coating for a 25-cm Whatman c18 column was 29 mg. For studies with the c8 quaternary salt, it was necessary to add the salt to the moblile phase; for a mo1.L-l concentration in the eluent, the retention times observed were similar to those with colunins coated with the larger molecular-weight salts. Changes in the column capacities (weight of sorbed quarternary salt) had pronounced effects on the relative retention of singly and doubly charged ions but only a minor effect within groups of ions of the same charge; this
-
Flgure 5. Column efficiency study for Cia column previously coated with tetraoctyiammonium iodide. Experimental conditions: Whatman column, 25 cm, 10 pm packing, coated with 90 mL of 5.8 X lo-' mo1.L-' tetraoctylammonium Iodide in 58 % ( v h ) methanol-42 % H 2 0 eluent, 0.2 mol-L-' NaH,PQ, and 0.039 mo1.L-i Na3P04(pH 6.4); sample, 20 pL of a solution containing 50-200 p9L-l of the anions; detection, UV absorbance at 215 nm.
0.027 ABSORBANCE UNITS
c 1 t
1
J , L l 400
RETENTION TIME ($1
Figure 6. Separation of anions on Cia column previously equilibrated with 200 mL of lo-, mol-L-l trldodecyimethylammonium iodide. Experimental conditions: coatlng solution, 75 % ( v h ) methanol-25 % H,O flow rate, 1 mL-min-'; numbers in brackets are HETP values in mm; other conditions, as for Figure 5. is the expected behavior for ion exchange separations. All of the column systems investigated had good column efficiencies and the data shown in Figure 5 are representative of these systems. The column efficiencieswere similar to those obtained for the cation systems described above; any large diferences between the HETP curves of the two systems were mainly due to differences between the column efficiencies of the uncoated reversed phases. A typical separation obtained with one of these quaternary salt systems is shown in Figure 6, and this chromatogram illustrates the rapid separations that are possible with these columns. All of the studies with columns coated with the CZ5,C32, and C37salts were in aqueous mobile phases which did not contain any of the quaternary salt. The results of initial studies with these column systems suggested that these sorbed modifiers were not as stable as the Cz0sulfate system. Consequently the long-term stability of these coated columns was studied. Figure 7 shows the changes observed in retention time and column efficiency for two anions, NO3- and I- as a function of total volume of eluate passed through a column coated with a C37quaternary salt; the results for I- represent the worst situation for the anions studied and the data for NO3-are typical of those for most of the other anions studied. These data were collected over a 2-week period and some of the fluctuations observed were due to large changes in the
ANALYTICAL CHEMISTRY, VOL. 54, NO. 9, AUGUST 1982
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1
0
no
.
I
I
5
10
15
VOLUME OF ELUENT (Ll
Flgure 7. Stability of sorbed trldodecylmethylammonium iodide on C,, bonded phase. Experlmental condltlons are as for Figure 5.
ambient temperture. The retention times were relatively constant, less so for I-, up to about 10 L of eluent, which represents 3 to 4 weeks of continuous use at 1 rnLemin-l. There was an initial rapid improvement in column efficiency, particularly for I-, followed by stable values up to 14 L. It appears that some reorientation of the bulky C3, quaternary salt took place initially which resulted in more efficient mass transfer. This effect did not appear to be as important for the lower molecular weight quaternary salts. Beyond 14 L there was a noticeable drop in column efficiency and retention time and a change in peak shape. When the columns were recoated the same poor peak shapes (and short retention times) were observed. Because these results suggested that some of the C18 bonded phase may have been removed, two of these columns were recoated "in situ" (17) with octadecyltrichlorosilane. Tests with these recoated columns showed that retention times for neutral organic compounds were similar to those found by the manufacturer, but peak shapes were poor and similar to those obtained for anions after 14 L of eluent. The column exhibiting the poorest peak shape was dismantled and a large void and a channel were found; these defects were likely responsible for the poor peak shape. The onset of these column defects may be related to poor packing structure, or to a slow deterioration of the reversed phase due to removal of the CISphase. If the latter is important then the use of buffers other than phosphate might prolong column life (18). Anion Exchange Systems-Styrenedivinylbenzene Phase. Unlike C18 bonded phases, styrenedivinylbenzene (SDVB) resins have not been widely applied to reversed-phase separations; however, the superior pH stability of the SDVB resins should permit the use of acidic and basic eluents, an important feature for anion determination (and cation) if extreme pH eluents are used. The quaternary salts were retained more strongly by the SDVB phase than by bonded phases. Good anion separations were obtained in the presence of the Cs quaternary salt (0.01 mo1.L-l in the eluent), but with larger molecular weight quarternary salts, which were coated onto the column by procedures similar to that used for the C18 bonded phases described above, the column efficiencies were poor unless small amounts of acetonitrile were added to the mobile phase. This transition from large to small HETP values was quite abrupt; for the C16 quarternary salt the transition took place in the 5-10% (v/v) acetonitrile composition range. The poor chromatography in pure aqueous eluents may be due in part to poor wetting of the surface of the support. Increased concentrations of acetonitrile above that required for good chromatography did not have any further appreciable effect. Methanol was significantly less effective than acetonitrile, even when large concentrations (50%)were used. Small concen-
-
RETENTION TIME Id
Figure 8. Separtion of anions on styrenedivinyibenzene reversed mo1-L-l trioctyimethylammoniumhydroxide In 1:l phase: curve A, acetonitrile-H,O; curve B, 1O3 moi-L-' tetrabutylammonium hydroxide in 15% (v/v) acetonitrile-85 % H20. Experimental conditions: flow rate, 1 mL.min-'; sample, 20 pL of a solution containing 9-50 pg-mL-' of each anion; detection, UV absorbance at 215 nm; numbers in
brackets are HETP values in mm.
trations of nonionic surfactants were only partially successful in improving the wetting of the support. The requirement for a partially nonaqueous eluent is a slight disadvantage relative to C18 bonded phases, because this will increase the solubility of the modifier in the eluent and necessitate the addition of small concentrations of the modifier to the mobile phase. Two chromatograms, which are representative of the separations obtained with the SDVB phase, are shown in Figure 8. These separations and the HETP values that are also given for each peak show that the column selectivity and efficiency are similar to those obtained with the c18 bonded phase (Figures 5 and 6). The SDVB column was used for over 4 months with a variety of eluents and modifiers without any observable loss in column behavior. This combination of stability and efficiency makes the SDVB phase an attractive choice for anion separations. Anion Detection. Absorption of UV light was used to monitor the test solutes used in these studies because it is sensitive and simple to apply; however, its selectivity is often a disadvantage and thus cannot be used to detect such important anions as C1-, P, SO-,: and PO:-. Conductivity can also be used for the detection of ionic species and gives a response for a much wider range of inorganic species; however, if it is to be used for the detection of ions in the pLg.mL-' and ng-mL-l ranges, the choices of eluent and column are limited. The detection limit obtainable with a conductivity monitor depends on two main experimental factors. The first, and normally less important factor, is the magnitude of the background signal which has a direct influence on the magnitude of the peak-to-peak base line noise and base line stability. The use of combination of low-capacitycolumna with low concentrations of eluents (2-7) helps reduce the importance of this factor. Because the capacity of the dynamically coated columns can be easily varied, they are ideally suited for such applications. The second and most important factor determining the sensitivity of conductivity detection is the magnitude of the difference between the equivalent conductivities of the eluent anion and the sample anion; each equivalent of the sample anion desorbed from the column on elution will replace an equivalent of the eluent anion which will be sorbed onto the exchange sites vacated by the sample anions. Consequently the eluent anion should have a low equivalent conductivity relative to that of the anion to be determined; improper choice can give zero or negative re-
ANALYTICAL CHEMISTRY, VOL. 54, NO. 9, AUGUST 1982
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Table 11. Repeatability of Anion Determinationsa
a
concn, fig.rnL-’
no. of samples Io; 124 7 4.1 8 NO; 34 7 8 1.1 NO; 34 7 8 1.1 141 7 8 1.4 Experimental conditions as for Figure 8. anion
re1 std dev in peak area, % 1 5 1 4 2
7 0.6 3
CONCENTRATION I pg m i - ’ )
Figure 9. Calibration curves for anion determination with styrenedivinylbenzene reversed phase. Experimental conditions: 2 X mo1.L-‘ tetrabutylammonium lhydroxlde, other conditions as for curve B, Figure 8.
sponse. We have used conductivity detection with both of the reversed phases used in this study and are presently investigating a variety of molbie phase compositions. The anions separated include F-,C1-, I-, Br-, NO3-,NO2-,SO4”, SO3”, and Po43-.The sensitivities obtainable can be illustrated by the results obtained for !3042which gives an intermediate response. With an aqueous (5% acetonitrile) mobile phase containing 5 x moEI,-’ each of tetrabutylammonium hydroxide and salicylic acid the detection limit (2X base line noise) for the 502-peak (K’ 5,HETP = 0.10mm) was 8 ng or 100 ng.mL-l for a 50-pL sample separated on a styrenedivinylbenzene phase. Suppressor column systems (1, 10) could possibly be used with these columns to improve sensitivity, but for the C18 bonded phases, the eluent must not be basic or rapid deterioration of the phase will occur. Basic eluents are compatible with the styrenedivinylbenrenephase and it should be possible to design a dynamically prepared exchange system with this phase that could be used with conventional carbonate eluents and cation suppressor columns. Supressor column systems have two main disadvantages however; they require periodic regeneration and degrade peak resolution. A continuous suppressor system consisting of a hollow-fiber ion exchanger has been studied (19) but the band spreading by these fiber8 must be reduced before they can be applied to HPLC separations. An alternate approach is to use a hydrophobic quaternary salt in the hydroxide form and add a weak organic acid at the column exit. The products from the reaction of the acid and the quaternary salt will be water and a hydrophobic ion pair. Studies of such systems as tetrabutylammonium hydroxide and phthalic acid showed that this approach improved sensitivities by 10- to 100-fold. This approach does not degrade column resolution and is a continuous process. Studies on the effects of eluent composition, structure of the quaternary salt, and structure of the acid on detection limits for this system are in progress.
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Quantitative Analysis. Figure 9 shows calibration curves obtained for four of the test anions separated on the SDVB support in the presence of tetrabutylammonium hydroxide; similar calibration plots were also obtained with the phosphate form of the modifier. The data in Table I1 show that good reproducibilities can be obtained in the low pg.mL-l range with a UV detector. This system has been applied to the determination of NO< and N O 1 (for the evaluation of processes being developed for the removal of nitrogen oxides from gas streams) in sample media such as deionized water, 6 m0l.L-1 ammonia, and 20% hydrogen peroxide. Problems such as those described for conventional ion chromatography (20) were not observed.
LITERATURE CITED (1) Small, H.; Stevens, T. S.; Bauman, W. C. Anal. Chem. 1975, 4 7 , 1801-1809. (2) Gjerde, D. T.; Frltz, J. S.; Schmuckler, G. J. Chromafogr. 1979, 786, 509-519. (3) Gjerde, D. T.; Fritz, J. S.; Schmuckler, G. J. Chromafogr. 1980, 787, 35-45. (4) Roberts, K. M.; GJerde, D. T.; Frltz, J. S. Anal. Chem. 1981, 53. 1691-1695. (5) Gjerde. D. T.; Frltz, J. S. Anal. Chem. 1981, 53, 2324-2327. (6) Harrlson, K.; Burge, D. Paper 301, 1979 Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy.
(7) Glrard, J. E.; Glatz, J. A. Am. Lab. (Fairfield, Conn.) 1981, 13 (lo), 26-35. (8) Green, L. W.; Huiland, J. S.; Sparkes, L. F., submitted for publication in J . Chromafogr. Scl. (9) Stevens, T. S.;Small, H. J. Li9. Chromatogr. 1978, 7 , 123-132. (10) Molnar. I.; Knauer, H.; Wiik, D. J. Chromafogr. 1980, 207, 225-240. (11) Reeve, R. N. J. Chromafogr. 1979, 177, 393-397. (12) Kirkland, J. J.; Yau, W. W.; Stoklosa, H. J.; Diiks, C. H., Jr. J. ChromtOgr. SCi. 1977, 75, 303-316. (13) Cassldy. R. M.; Elchuk, S. Anal. Chem. 1979, 5 7 , 1434-1438. (14) Cassidy, R. M.; Elchuk, S. J. Chromatogr. Scl. 1980, 4 , 217‘-223. (15) Cassidy, R. M.; Elchuk, S. J. Ll9. Chmmatogr. 1981, 4 , 379-398. (16) McCormic, K.; Karger, B. L. Anal. Chem. 1980, 5 2 , 2249-2257. (17) Giipin, R. K.; Camlllo, 0.J.; Janicki, C. A. J . Chromafogr. 1976, 121, 13-22. (18) Synder, L. R.; Kirkland, J. J. “Introduction to Modern Llquld Chromatography”, 2nd ed.; Wliey: New York, 1979;p 296. (19) Stevens, T. S.; Davis, J. C.; Small, H. Anal. Chem. 1981, 5 3 , 1488-1492. (20) Koch, W. F. Anal. Chem. 1979, 5 7 , 1571-1573.
RWEIVED for review March 19,1982. Accepted April 30,1982. This is Report AECL-7724.
