Anal. Chem. 1992. 6 4 , 853-862 (5) Raymer, J.; Pellizreri. E. Anal. Chem. 1987, 5 9 , 1043-1048. (6) Raymr, J.; Pelllrzarl, E.; Cooper, S. Anal. Chem. 1867, 5 9 , 2069-2073. (7) MCNaHy, M. E. P.; WhWkn. J. R. J . C h r O M t m . 1888. 435, 63-71. (8) Stahl, E.; QuMn, K.; Glatz, A.; Gerard, D.; Rau. G. Ber. Bunsen-Ges. PnyS. Chem. 1984, 88, 900-907. (9) B o d a n d ConjqgttedPestb& RWws; Kaufman, D. D., Still, G. G., Paulson, G. D., Bandal. S. K., Eds.; ACS Symposium Series 29; American Chemical Soclety: Washington, DC, 1976; Chapter 3. (10) ~ d e n d C o n l u g e W P e s t b U sR e s k f w s ; Kaufman. D. D.. Still, G. G.. Paulson, G. D., Bendal. S. K., Eds.; ACS Symposium Series 29; American Chemical Soclety: Washington. DC, 1976; Chapter 5. (11) Brlstol, D.; cook. L.; Koterba, M.; Nelson, D. J . Agrlc. food Chem. 1882, 30, 137-143.
853
(12) Norwitz, 0. Anal. Chem. 1968, 58, 639-641. (13) Narang. A. S.; Vernoy, C. A.; Eadon, G. A. J . Assoc. Off. Anal. Chem. 1983, 86, 1330-1334. (14) Chesney, D. J.; TaHman. D. E.; Peckrul, A. A,; Cook, L. W.; Fleeker, J. R. Anal. Chlm. Acta 1987, 197, 159-167. (15) Van Leer, R. A.; Paulakis. M. E. J . Chem. €ng. Data 1980. 2 5 , 257-259. (16) Decedue. C. J.; Unruh, W. P. BioTechniques 1984. 2 . 78-81. (17) Thomson, C.; Chesney. D. J . Chromatogr. 1991, 543, 157-194. (18) King, J. W. J . Chrometcg. Sci. 1989, 27, 355-364. (19) Khan, S. U. J . A M . FoodChem. 1980. 28, 1096-1098.
RECW
for review July 18,1991. Accepted January 15,1992.
The Role of Lewis Acid-Base Processes in Ligand-Exchange Chromatography of Benzoic Acid Derivatives on Zirconium Oxide John A. Blackwell* Group Analytical Laboratory, Specialty Adhesives and Chemicals Division, 3M Company, 236-2B-11,3M Center, St. Paul, Minnesota 55144-1000
Peter W. Carr Department of Chemistry and Institute for Advanced Studies in Bioprocess Technology, University of Minnesota, 207 Pleasant Street SE, Minneapolis, Minnesota 55455
Porous mkropartlculate zlrconlun oxlde drows very dlfferent selectlvlties and pH dependencks for the separatlon of benrolc acld ckrlvatlves than do conventlonal bonded-phase anlon-exchange supports. Thls results from a very dgnlflcant Ilgand-exchage contrlbutlon to the retentlon of hard Lewk ba-8 on the “face of trarttlonmetal oxlde supports. We have found that the capaclty factors of a wlde varlety of derlvatlves of benrolc acld are closely correlated wlth thelr Bron8t.d acldltles. The eluent pH k also a crltlcal factor In detennhlng the magnttude of the capadty factor, but lt does not have much Muence on chromatoQaphk selecthrlty. The dlfferentlal sdectlvlty of this phase In comparkon to conventbnal polvmerlc and bondebphase anbn exchangers can be attributed to complexallon and steric effects whlch profoundly alter the elutlon patterns of certaln solutes.
INTRODUCTION While the literature on inorganic ion exchangers is quite broad,l application of these materials to high-performance liquid chromatography is rare. In this work, we report on the uae of microparticulate porous zirconium oxide as a stationary phase for the chromatographic separation of carboxylic acids. Previously, we described the use of polymer-coated zirconia as a support for reversed-phase separations and noted the very strong binding of carboxylic acids to the underlying zirconia surface? Reports have appeared on the use of microparticulate alumina as a stationary phase for the ion-exchange separation of inorganic however, the role of Lewis acid-base and ligand-exchange processes was not recognized.
* To whom correspondence should be addressed. 0003-2700/92/0364-0853$03.00/0
In comparison to ligand-exchange chromatography, ionexchange chromatography is relatively straightforward. Since the present work pertains primarily to the separation of anions, we will briefly consider the retention of a set of simple anions (denoted A;) on a conventional strong base anionexchange support. The anion-exchange process can be written as
R+:X- + A- = R+:A- + X-
(1)
Even if a strong anion exchanger is used, the eluent pH can significantlyalter retention. For simplicity, let us also assume that X-,the counterion to the fixed charge, R+, is a strong electrolyte, but that HA, the Bronsted conjugate of A-, is a weak acid. If we assume that only Coulombic interactions are significant, that is, we ignore secondary retention processes such as hydrophobic and hydrogen-bonding forces, then only A-, and not HA, will be retained. Clearly, retention will decrease as the pH is decreased since a greater fraction of the sample will exist in the HA form. Similarly, we expect that solute retention will decrease as the pK, of the solute increases, since at a fixed eluent pH a greater fraction of a solute with a higher pK, will exist in the unretained (HA) form. In this work, we will show why these simple ideas are not applicable to retention in ligand-exchange chromatography on metal oxide surfaces due to the strong contributions of Lewis acid-base interactions to retention. The use of inorganic (metal oxide) ligand exchangers for the separation of low molecular weight solutes, such as carboxylic acids, is relatively unexplored. Stumm et al. demonstrated the ligand-exchange properties of hydrous oxides and showed that the strength of the interaction of coordinating solutes was a function of their Lewis basicity, geometry, and for those solutes which are also weak electrolytes, Bronsted a ~ i d i t y . ~Ligand exchange takes place at the Lewis acid 0 1992 American Chemical Society
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ANALYTICAL CHEMISTRY, VOL. 64, NO. 8, APRIL 15, 1992
(cationic metal) sites of the metal oxide surface. A good correlation was found between experimentally determined surface formation constants and solution formation cons t a n t ~ .The ~ ~ strongest ~ interactions take place when more than one of the donor groups of a multidentate solute (a chelon) simultaneously interacts with a single Lewis acid surface site. However, Stumm’s studies were carried out statically and his results have never been tested under the dynamic conditions found in chromatography. Regazzoni et al. used data on the adsorption of various anions on zirconia, magnetite and hematite to develop a model of ligand exchange which involves both site binding and surface c o ~ r d i n a t i o nThis . ~ ~ ~model accurately describes the adsorption characteristics of metal oxides toward cations and anions as well as both complexing and noncomplexing solutes. It predicts the surface potential changes caused by sorption of such species. The following equations were developed to rationalize the electrophoreticbehavior of hematite in fluoride and formate media:7
+ H+ = M(H,O)Z+ M(OH)(H20) = M(OH)2- + H+
M(OH)(H,O)
M(OH)z- + K+
M(0H)Z- K+
M(OH)(HzO) + L1- = M(H,O)L,+ OH-
(2)
(3)
(5) (6)
where L1 and M represent a Lewis base and the metal oxide metal ion, respectively. In contrast to silica, transition-metal oxide surfaces are amphiprotic, that is, they can act as both acids and bases and thus can serve as both anion and cation exchangers. Process 2 corresponds to the protonation of an uncharged surface Bronsted base site. On zirconia the resulting conjugate acid has a pK, of about 4.0a9Process 3 corresponds to the loss of a proton from a second type of site to form an negatively charged site. Experimental data indicate a pK, of approximately 8.L9 The remaining reactions provide the basis for simple anion and cation exchange (processes 4 and 5 , respectively) and for ligand exchange of L1- for a bound hydroxide group or for bound water (processes 6 and 7 , respectively). These equations were developed by Regazzoni et al. by combining the site binding and surface coordination models of adsorption at the metal oxide/solution interface. They are fully consistent with the adsorptive behavior of the metal oxide. Processes 2 and 4 account for the anion-exchangeproperties of zirconia at low pH for noncoordinating anions, such as chloride, nitrate, and perchlorate. AE the pH is increased, the anion-exchange capacity of the material decreases,as predicted by the shift to the left in process 2, and retention of noncoordinating anions should decrease. At higher pH, process 5 accounts for the cation-exchange properties of the support toward noncoordinating cations. Between these two pH limits the anion- and cation-exchange capacity of the suIface varies in accord with the solution pH. Processes 6 and 7 describe the displacement reactions at the surface when a Lewis base eluent is used. On the basis of relative formation constants, we expect process 7 to be more important than process 6 since hydroxide ions are much more tightly bound by zirconia than are water molecules. However, the displacement is governed by mass action and the eluent pH will determine which process is more significant. As observed in prior pH studieslOJ1of the adsorption of fluoride ion, which is a very strong Lewis base toward zirconia, water is much more readily displaced from the surface than is hy-
droxide ion. Lewis bases which are weaker than fluoride will displace even less hydroxide relative to water. We believe that Regazzoni‘s model can be used to describe the retention of a variety of species on zirconia, including Lewis base solutes, in Lewis base containing eluents by the following additional processes: M(OH)L1- + L2- = M(0H)Lz-
+ L1-
+ Lz- = M(0H)Lz- + H2O M(OH)(HzO) + Lz- = M(H2O)LZ + OHM(OH)(H,O)
(8) (9)
(10) where Lp- represents a solute Lewis base. These additional processes take into account the exchange of an eluent Lewis base (L1J for a solute base (L2-). Processes 8-10 account for the displacement of an eluent (buffer) Lewis base, a bound water molecule, and a bound hydroxide by a solute Lewis base, respectively. The above set of equilibria can be used to describe the chromatographic characteristics of many different solutes in a wide variety of eluents that contain competing Lewis bases. On the basis of the above theoretical picture of the static surface properties, it is necessary to determine the experimental parameters which affect the ligand-exchange properties of metal oxides in a flowing system. The thermodynamic and kinetic parameters for ligand exchange must be determined in order to make ligand exchange on supports such as zirconium oxide possible. To accomplish this,the chromatographic properties of a series of benzoic acid derivatives on zirconium oxide were examined in order to determine whether ligandexchange chromatography is feasible. The results are compared to data obtained using a conventional strong anion exchanger to determine the differences in chromatographic behavior caused by the ligand-exchange interaction. EXPERIMENTAL SECTION Chemicals. 2-(N-Cyclohexylamino)ethanesulfonic acid (CHES),N-(2-hydroxyethyl)piperazine-A”-3-propanesulfonicacid (EPPS),N-(2-hydroxyethyl)piperazine-N’-2-ethanesulfonicacid acid (MES), 34N(HEPES), 2-(A’-morpholino)ethanesulfonic morpho1ino)propanesulfonicacid (MOPS),piperazine-A’,”-bis(2-ethanesulfonic acid) (PIPES), and 3-[N-[tris(hydroxymethyl)methyl]amino]propanesulfonic acid (TAPS) were obtained from Sigma (St. Louis, MO). Hydrochloric acid and HPLC grade 2-propanol were obtained from Fisher Scientific (FairLawn, NJ). Sodium hydroxide was obtained as a 50% solution from Curtin Matheson Scientific (Houston, TX). Acetic acid was obtained from EM Science (Cherry Hill, NJ). All other chemicals were obtained from commercial sources and were reagent grade or better. Water used in these studies was prepared by passing housedeionized water through a Barnstead NanoPure water system with an additional organic-freecartridge and a 0.2-pm final filter. The water was subsequently boiled for 5 min and then cooled to room temperature immediately prior to the use to remove dissolved carbon dioxide. Chromatographic Supports. The porous zirconium oxide particles were provided by the Ceramic Technology Center of the 3M Co. and were described earlier.2JO-16 The particles used in this investigation had a nominal diameter of 5.3 f 1.3 pm, an average pore diameter of 308 A by mercury porosimetry, and an average BET surface area of 32.5 m2/g. The particles were initially pretreated in order to remove as many of the manufacturing impurities as possible, as described earlier.1°J1J6 Chromatographic columns were prepared in 50- X 4.6-mm column blanks fitted with 1/4-in.Parker end fittings. Titanium screens with 2-gm mesh were used instead of frits to minimize any potential extraneous metal ion contamination from the frits. Columns were packed by the upward slurry technique using 2-propanol as the solvent. Packing pressure was 4500 psi (300 atm). Following the packing procedure, all columns were flushed with water to displace all of the packing solvent prior to introduction of the buffer.
ANALYTICAL CHEMISTRY, VOL. 64, NO. 8, APRIL 15, 1992
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Table I. pH Dependence of t h e Capacity Factors of Benzoic Acids on Zirconium Oxide Supportsa PH derivative
PK,
4.8
6.1
6.8
7.2
7.5
8.0
8.4
9.3
4-nitro 4-cyano 4-formyl 4-chloro 4-bromo 4-iOdO benzoic acid 4-acetamido 4-ethyl 4-isopropyl 4-methyl 4-hydroxy 4-ethoxy 4-amino 4-dimethylamino 2-iodo 2- hydroxy 2-phenoxy 2-methoxy 2-acetyl 2-ethoxy 2-amino 2-carboxy
3.44 3.55 3.76 3.99 3.99 4.00 4.20 4.28 4.35 4.36 4.36 4.58 4.80 4.85 5.00 2.86 2.98 3.53 4.09 4.13 4.21 4.79 5.41
4.4 5.2 6.5 11.3 11.8 13.4 11.7 21.0 22.7 19.8 20.4 27.3 26.3 41.0 57.2 0.9 eno
31.9 31.0 39.4 eno eno eno 76.5 eno eno eno eno eno eno eno eno 9.8 eno 23.4 44.4 25.7 42.0 eno eno
4.6 4.3 5.4
2.1
0.0
0.2
3.7 3.4 3.6 3.5 5.9 4.3 4.1 8.0 8.4 5.7 12.8 26.2 0.5 eno 0.8
0.5 0.3 0.4 0.6 0.6 0.6 0.6 0.8 0.7 0.7 1.8 1.4 1.0 1.7 4.4 0.1 eno 0.1 0.3 0.2 0.2 3.7 eno
0.3
11.8 12.2
1.8 1.3 0.9 3.2 2.8
0.0 0.0 0.0 0.0 0.0 0.0
2.7
4.9 1.4 2.7 30.6 eno
1.7 2.2
13.7 10.3 21.7 18.1 14.0 21.3 24.7 24.4 50.5 82.7 1.3 eno 2.8 7.6 3.3 5.4 67.7 eno
2.2 1.2
1.3 21.1 eno
2.1
1.4 5.1 3.1 2.1
6.5 3.1 4.4 11.4 20.1 0.3 eno 0.3 1.0 1.0 1.2 18.1 eno
0.3 0.4 0.5 0.6 0.4 0.8 0.8 0.7 1.4 1.4 1.0 1.6 4.2 0.0 eno 0.1 0.3 0.1 0.1 2.8 eno
0.1 0.0 0.0 0.2 0.1 0.1 0.2 0.4 0.0 26.9 0.0 0.0 0.0 0.0
0.4 3.0
aEluent was 100 mM acetate at pH 4.8, MES at pH 6.1, PIPES at pH 6.8, MOPS at pH 7.2, HEPES at pH 7.5, EPPS at pH 8.0, TAPS at pH 8.4, or CHES at pH 9.3. The flow rate was 1.0 mL/min at 35 "C. Injections were 10-pL volumes of 10 mM solutions. Detection was at 230, 254, and 280 nm. eno = elution not observed (k' > 100). Anion-exchange chromatography was performed on a 150- X 4 . 6 " Spherisorb 5 SAX column containing 5-pm particles. This column was obtained from Phenomenex (Torrance, CA) and was flushed with freshly boiled water prior to equilibration with the eluting buffers. Chromatographic Systems. Chromatographic studies were carried out on a Hewlett Packard (Avondale, PA) Model 1090M liquid chromatograph with a DR5 ternary solvent delivery system and a diode array detector. The optional expanded pH range kit as well as ultrahigh molecular weight polyethylene piston seals (UPC-10) obtained from Bal Seal Engineering (Santa Ana, CA) were installed. Data were processed using a Hewlett Packard 9000/Series 300 computer outfitted with ChemStation software. This system was outiftted with a 50- X 4.6" column filled with 1G20-pm zirconia particles. This guard column was placed before the injection valve to scavenge any metal ion contaminants in the buffer. Retention Studies. The capacity factors and reduced plate height data were obtained by choosing benzoic acid derivatives which spanned a large pKa range and had functional groups of widely varying Lewis and Bronsted acidity and basicity. These solutes were diasolved in water to make a final solute concentration of approximately 10 mM. Many solutions were saturated at concentrations lower than this target concentration and thus were used as the saturated solution. Because of the small injection volumes involved (10 pL), sample pHs were not standardized. In addition to the tabulated results, other derivatives (mainly meta derivatives) were chromatographed as well. Buffers consisted of 100 mM solutions of acetate a t p H 4.8, MES a t p H 6.1, P I P E S a t p H 6.8, MOPS a t p H 7.2, HEPES a t p H 7.5, EPS a t p H 8.0, TAPS a t p H 8.4, or CHES at p H 9.3. Solutions above p H 7.5 were not used on the Spherisorb 5 SAX column due to the instability of silica-based packing materials in alkaline buffers. Columns were equilibrated with 200 mL of eluting buffer a t a flow rate of 1.0 mL/min a t 35 "C prior to beginning the study. Injections (10 pL) of each solute.were made, and elution was monitored at 230,254, and 280 nm using a diode array detector. Capacity factor measurements were made using the peak maximum. Asymmetry factors for solutes on the zirconium oxide phase, as determined by the ratio of a / b a t 10% of the peak height, were fairly constant a t approximately 3 for all solutes except those which were strongly retained (k'> 30). These solutes showed much higher asymmetry factors with some approaching 6. Peaks eluting from the Spherisorb phase typically
showed asymmetry factors of 1.5-2.2. Reduced plate heights were determined using the width a t half-height method. Due to the asymmetry of the peaks, these reduced plate heights deviate from those calculated from peak moments. This error was disregarded since nearly all the peaks were of similar shape (hence similar error from true plate heights) and the relative efficiencies were important rather than the absolute efficiencies. According to the analyses using a particular Lewis base eluent, the zirconium oxide columns were flushed with 45 mL of 0.1 M sodium hydroxide to strip any "irreversibly" bound solutes. This was followed by a 50-mL flush with water to remove excess hydroxide from the column. Previous studies of the adsorption and desorption of fluoride indicate that this treatment should suffice to remove any sorbed Lewis base.1°J6 Reproducibility was better than 5% for capacity factors and 7% for reduced plate heights despite repeated base stripping. The effect of p H on the retention of benzoic acid derivatives and other probe solutes was also determined using citric acid as the Lewis base buffer. Isocratic elution of the solutes was performed using solutions containing 0.2 M sodium chloride in 20 mM citric acid at a variety of pHs. The pHs tested spanned the range from one-half p H unit below the lowest pK, of citric acid to one p H unit above the highest pKa of citric acid. Injections (10pL) of the test solutes (10 mM in water) were used. The flow rate was 0.5 mL/min at 35 "C with detection a t 254 and 280 nm. Capacity factors were determined from the location of the peak maximum and the reduced plate heights calculated using a particle size of 5.3 pm.
RESULTS AND DISCUSSION Solute Structural Effects on Retention. T h e two most important factors controlling t h e retention of both coordinating and noncoordinating species on zirconia supports are the type of eluent Lewis base and t h e eluent's pH. Table I gives the capacity factors of 23 ortho- and para-substituted benzoic acid derivatives on zirconia as a function of p H using a variety of buffers to control the pH. Data at all b u t one pH were obtained in amino sulfonate buffers. This type of buffer was chosen because sulfonates are only moderately strong Lewis bases toward zirconia1°J1J6 and their use allowed us t o cover a wide range in pH without greatly altering t h e chemistry of the eluent's interaction with zirconia. T h e pH of these buffers is controlled by differences in the pK,'s
of t h e con-
858
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1
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Table 11. Regression Analysis of Retention of p-Benzoic Acids vs pK, on Zirconia pH
slope"
std dev*
Y intc
std de@
rZe
nf
4.8 6.8 7.2 7.5 8.0 8.4 9.3 overall
0.67 0.73 0.62 0.62 0.56 0.71 0.80 0.67
0.04 0.07 0.08 0.13 0.09 0.08 0.19 0.10
-1.65 -1.92 -1.93 -2.14 -2.46 -3.19 -4.75
0.07 0.11 0.14 0.22 0.16 0.13 0.33
0.95 0.90 0.82 0.64 0.74 0.86 0.57
15 15 15 15 15 15 15
"Slope of log,, k' vs pK, for p-benzoic acids. *Standard deviation of the slope. CInterceptof the regression. dStandard deviation of the intercept. e Correlation coefficient. f Number of pointa.
U
LL
x +
.-0 U
Q
A
Table 111. pH Dependence of the Capacity Factors of Benzoic Acids on Spherisorb 5 SAXa
U
DH
0
GO'
I Y
I
1
I
,
3 ;5 350 3 75 400 425 4 5C 4 75 5C0
pKa Flguro 1. Plot of log k'of p-benzoic acid derhratives vs their pK, on zirconium oxide in various buffers: (0)acetate at pH 4.8; (0)PIPES at pH 6.8; (V)MOPS at pH 7.2; (V)HEES at pH 7.5; ( 0 )EPPS at pH 8.0; (a)TAPS at pH 8.4; (A)CHES at pH 9.3. All conditions are as
in Table I. SoiM lines are least-squares fits.
stituent amino groups. While amino groups are classed as hard Lewis bases, the amino groups in the amino sulfonate buffers are highly hindered. Other ~ o r k ' ~ indicates J~ that the amino sulfonate buffers are very weak eluents on zirconia. For our present purposes we believe that the interactions between the amino groups of the amino sulfonate buffer and zirconia are, to a first approximation, of no importance. A number of trends are clearly evident in Table I. First, the benzoic acids are well retained in the low-pH (6.1) amino sulfonate eluent. Second, on zirconia, those benzoic acids that have hard Lewis base active substituents (e.g. COOH, OH, and unprotonated primary aliphatic NH2groups) ortho to the carboxylic acid moiety of the benzoic acid have extraordinarily high capacity factors compared to the retention of the corresponding para isomer. This heightened retention is not observed on a conventional strong acid ion exchanger (see below). This second observation strongly supports our view that t h e formation of coordination complexes between Lewis base solutes and Lewis acid surface sites can be a very important contributor to retention on zirconia. Analogously, Stumm4 found that bidentate solutes have anomalously large sorption coefficients (Kd) on metal oxides and took this as proof of the existence of ligand-exchange interactions. Similarly, in his classic study of adsorption of alumina, SnyderlsJg found that the retention of ortho dicarboxylic acids and other ortho di-Lewis base substituted solutes was much greater than that predicted by the sum of the individual contributions. Conversely, when only one Lewis base group is present in an ortho isomer, then, due to steric hindrance, the total adsorption strength is less than the s u m
derivative
4.8
6.1
6.8
7.2
7.5
4-nitro 4-CymO 4-formyl 4-chloro 4-bromo 4-iOdO benzoic acid 4-acetamido 4-ethyl 4-isopropyl 4-methyl 4-hydroxy 4-ethoxy 4-amino 4-dimethylamino 2-iodo 2-hyd10xy 2-phenoxy 2-methoxy 2-acetyl 2-ethoxy 2-amino 2-c~boxy
2.8 2.0 2.3 4.7 5.5 6.7 10.3 2.7 6.7 3.7 5.1 1.7 5.0 2.0 6.3 4.5 4.2 10.3 3.5 3.3 4.2 2.5 7.4
0.5 0.4 0.5 0.8 0.8 0.9 0.6 0.7 1.0 0.7 1.0 0.4 1.0 0.6 1.7 0.7 0.6 1.2 0.7 0.7 0.8 0.7 0.8
0.2 0.2 0.2 0.3 0.3 0.3 0.3 0.3 0.4 0.3 0.3 0.2 0.4 0.3 0.6 0.3 0.3 0.4 0.3 0.3 0.3 0.3 0.2
0.3 0.3 0.2 0.5 0.6 0.6
0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.2 0.1 0.1 0.1 0.1 0.1 0.3 0.1 0.1 0.2 0.1 0.1 0.1 0.1 0.0
0.3
0.4 0.7 0.4 0.8 0.3 0.6 0.4 1.2 0.4 0.4 0.7 0.4 0.4 0.4 0.6 0.3
" See Table I for conditions. of the individual contributions. In a related study of adsorption on silica, Snyder showed that due to internal hydrogen bonding and steric factors, ortho isomers are less retained than are para i s ~ m e r s . ' ~ This J ~ occurs because silica does not have the Lewis acid sites necessary for ligand-exchange interactions, and thus it cannot show the type of chelation effect that takes place on transition-metal oxide surfaces. The effect of substituent type on retention was examined in more detail by plotting the decadic logarithm of the capacity factor vs the solute's pK, (see Figure 1). In order to avoid the chemical complexities introduced by the chelation and steric factors alluded to above, only the para-substituted derivatives were plotted. As shown in Figure 1, a series of roughly linear relationships are obtained in each buffer. Table I1 summarizes the regression data The average slope is +0.67 (fO.l). What is most striking about these data is the fact that in contrast to the behavior predicted for a simple strong base anion exchanger (see above), retention increases as the solute pK, increases. It should be noted that in the amino sulfonate buffers, where the pH is 6.1 or greater, all of the solutes are at least 95% ionized. Consequently, the correlation between the log k' and the pK, cannot be due to variations in the fraction of the solute in the protonated form. This interpretation was substantiated by examining the retention of a
ANALYTICAL CHEMISTRY, VOL. 64, NO. 8, APRIL 15, 1992 i 00
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c
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30
I
1
I
1
35
40
45
50
55
p K o (Solute)
Flgure 9. Plot of the retention of p-benzoic acid derivatives vs pK, at pH 4.6: (0)zirconium oxMe column; (0)Spherisorb SAX column. The eluent was 100 mM acetate with a flow rate of 1.0 mL/min at 35 OC.
series of benzoic acid derivatives on a silica-based bondedphase strong acid anion exchanger (see Table I11 and Figures 2 and 3). It is evident that the pKa has very little correlation with the capacity factor. Further, we note that ortho dicarboxylates and other solutes having two Lewis base active groups in the ortho position are not particularly strongly retained on the conventional column. Thus the correlation observed in Figure 1must be related to the chemistry of the interaction of the solute with the zirconia surface. The silicaand zirconia-based supports are compared in more detail below. We believe that the relationship between log k’and pK, occurs because there is a correlation between the Bronsted basicity of the anionic form of the solute and the Lewis basicity of the solute toward zirconium. In the following discussion one must keep in mind the fact that as the Bronsted acidity of HA weakens, the Bronsted basicity of the conjugate base increases, that is, the affiiity of A- for protons increases. The similarity of the slopes of log k’vs pKa a t all pHs shown in Figure 1demonstrates that the Bronsted acidity of the carboxylic acids is directly related to the Lewis basicity of the conjugate carboxylate anion. This is more easily reconciled
857
when one considers that the proton is the quinteseential, hard Lewis acid. Similarly, zirconium(IV) is an exceptionally strong hard Lewis acid. The affinity of the benzoate anion for the proton is directly measured by its Bronsted basicity. Since protons and zirconium ions are both hard Lewis acids, the correlation between proton affiiity and zirconium ion affinity should be high. If all of the changes in log k’ for the parasubstituted solutes are attributed to changes in the Lewis basicity of the solute then the common slope of 0.67 for plots of log k‘ vs the pKa indicates that there is a rather strong relationship between Bronsted basicity and Lewis basicity toward zirconia. Several points must be borne in mind before generalizing the above conclusion. First, the correlation between retention and pK, is not overwhelmingly strong, though it is certainly statistically real. However, a more detailed factor analysis of the data (results not given) for the para-substituted acids clearly revealed the existence of three distinct principal components related to the probe solute. Only one of these factors was strongly correlated with the solute pKa. It is likely that Lewis basicity toward zirconia depends on several factors, only one of which is gauged by the pK,. Second, the correlation is based on only one type of carboxylate group, that is, an aromatic carboxylate moiety. Under other conditions (not shown here) we have observed a 3-fold difference in retention and a 10-fold difference in the rate of desorption of phenylacetic acid, which has an aliphatic carboxyl group, relative to the retention of p-methylbenzoic acid despite the fact that both have identical pK,’s. While we believe that the solute Lewis basicity will be an important factor in determining retention, it is likely that aliphatic and aromatic carboxylates will follow different correlations. Eluent pH Effects on Retention. Again, in contrast to the behavior expected on a strong acid anion exchanger the data of Table I and Figure 1show that except for the results in acetate media, the capacity factors of all solutes decrease as the eluent pH increases. Similar pH-dependent adsorption properties have been observed in studies of other metal oxidea?*22 When we regressed the log k ’data for the individual solutes vs pH, using only the amino sulfonate buffers (see Table IV), we found that the mean slope was approximately -0.90 (h0.12). Examination of the data of Table I11 for the silica-basedstrong acid anion exchanger shows that k ‘is almost independent of pH except for those solutes which have protonatable amino groups. As stated above, all of the acids are virtually completely ionized over the entire range of pHs explored in the amino sulfonate buffers (pH > 6.1) and thus we expect no pH dependence based on changes in the degree of ionization of the solute. We conclude that the pH effect is primariiy due to processes 3,6, and 10. That is, the decrease in k ’is due to a decrease in the positive charge of the metal oxide surface by removal of protons and, more importantly, due to increased competition between hydroxide and the solute for occupying Lewis acid surface sites. This is in agreement with our previous study of the adsorption of fluoride on zirconia.1°J6 In that work, we showed that in neutral and slightly alkaline media hydroxide is able to displace fluoride from zirconia. This is very significant because fluoride is a far stronger Lewis base toward zirconia than are most carboxylates and thus hydroxide should be a stronger competitor with carboxylates than it is for fluoride. The closeness of the observed slope of log k’vs pH to -1 (-0.90 f 0.12) suggests that the Lewis basicities of the various amino sulfonates are very similar (however, see below). It should be noted that at all of the pHs used in this study the sulfonate moiety of the amino sulfonic acids was completely ionized. We have no strong evidence to suggest that hindered secondary and tertiary amino groups of the amino sulfonate
858
ANALYTICAL CHEMISTRY, VOL. 64, NO. 8, APRIL 15, 1992
Table IV. Regression Analysis of Retention of p-Benzoic Acids vs pH on Zirconia derivative
slope’
std de$
Y intc
std de@
r2e
nf
nitro cyano formyl chloro bromo iodo benzoic acid acetamido ethyl isopropyl methyl hydroxy ethoxy amino dimethylamino
-0.86 -0.96 -0.94 -0.94 -0.89 -0.87 -1.01 -0.91 -0.87 -0.84 -0.79 -0.89 -0.90 -0.93 -0.89
0.06 0.06 0.09 0.10 0.12 0.14 0.08 0.08 0.16 0.14 0.05 0.09 0.08 0.08 0.07
6.66 7.22 7.19 7.43 7.03 6.91 7.94 7.41 7.05 6.69 6.65 7.34 7.35 7.95 7.91
0.12 0.12 0.16 0.13 0.15 0.18 0.16 0.16 0.20 0.17 0.09 0.18 0.16 0.17 0.13
0.98 0.98 0.97 0.97 0.95 0.93 0.97 0.97 0.91 0.93 0.99 0.96 0.97 0.97 0.98
7 7 7 6 6 6 7 6 6 6 6 6 6 7 7
a Slope of log,, k’ vs pH for p-benzoic acids. * Standard deviation of the slope. Intercept of the regression. Standard deviation of the intercept. e Correlation coefficient. ’Number of data points.
3
I
1
shown that under the same elution conditions, phenylacetic acid is much more strongly retained than is benzeneeulfonic acid. Thus part of the change in retention between the acetate and sulfonate eluents is due to a shift in the relative contributions of processes 6-10 (K(acetate) > K(amino sulfonate)) and part is a pH effect on the solute and on the zirconia surface. The pH 8.4 TAPS buffer appears to be a slightly stronger eluent relative to the other amino sulfonate eluents. This effect may be the result of the tris(hydroxymethy1)aminomethane group on this buffer salt which acts to chelate the Lewis acid sites. In previous work on zirconium phosphate we observed that TRIS buffer, which also contains a tris(hydroxymethy1)aminogroup, interacted very strongly with zirc~nium.l~J~-~~ Figure 4 suggests that eq 11can be used to predict k’values for these benzoic acid derivatives, where C is a constant which differs for chemically distinct displacing buffers, that is, C is log k’ = 0.67pK, - pH C
Y
/
/
2
1
Iy
m
o
i
-1
+
-2 /
-3
-e
-6
-4
-2
c
( G 7 p K a - pH) Flgurr 4. Correlation of the capacity factors of p-benzoic acids on zlrconlum oxide wlth solute Bronsted addity and solutkn pH: (0)amino sulfonate buffers: (0)acetate buffer. Regressions are least-squares fits. Data are the same as In Figure 1.
buffers are able to coordinate to zirconia. In order to compare retention in all of the buffers, including the acetate buffer, the log,, k’values were “corrected”for their dependence on pH by plotting them against the function of pK, and pH shown in Figure 4. In doing so we assumed that the theoretical dependence on pH and pK, should have slopes of -1 and 0.67, respectively. While there is a great deal of scatter, it is clear, given the large number of solutes and conditions, that all of the data in the amino sulfonate buffers fall on one line and that the data in the acetate buffer are distinctly different. On the basis of this pH “corrected” retention plot (Figure 4),retention in the acetate-containing eluent is clearly much weaker than in amino sulfonate buffers. We believe that this results primarily from the more intensive competition between acetate and the solute rather than between a sulfonate and the solute for the surface Lewis acid sites. This is not surprising because simple carboxylates are stronger Lewis basea than are sulfonates. we have
the same for all amino sulfonate eluents but is a different number for an aliphatic carboxylate buffer. In the companion paper,l’ we will show that it is possible to generate an eluotropic scale based on the ability of a Lewis base to displace a variety of benzoic acid derivatives from the surface of zirconia. Solute and p H Dependence of the Column Efficiency. We must emphasize the fact that the peak widths and column efficiencies obtained in ligand-exchange chromatography on zirconia supporte are often quite poor in comparison to those observed in ion-exchange chromatography on conventional bounded-phase columns. On average, the reduced plate heights are much poorer than usually observed in a wellpacked column with particles having uniform meso pores. Chromatography of nonbinding, nonionic solutes gave reasonable reduced plate heights.l6B In this work, there are many cases in which the plate heights are so large that they cannot possibly be due to anything other than very slow chemical desorption of the solute from the stationary phase. This is substantiated by the fact that the plate heights are very eluent and solute dependent even under conditions such that the k’ valuea are very similar. Assuming that the chemical deaorption step is the rate-limiting step, Gidding’s work2‘ leads to the following relationship: 2uk’ kd = H,(1 + k’12 where h is the reduced plate height, k ’is the capacity factor, u is the linear velocity of the mobile phase, and kd is the rate
ANALYTICAL CHEMISTRY, VOL. 64, NO. 8, APRIL 15, 1992
859
Table V. pH Dependence of the Desorption Rate Constant for Benzoic Acids on Zirconium Oxide Supports" PH derivative
4.8
6.1
6.8
7.2
7.5
8.0
8.4
9.3
4-nitro 4-cyano 4-formyl 4-chloro 4-bromo 4-iOdO benzoic acid 4-acetamido 4-ethyl 4-isopropyl 4-methyl 4-hydroxy 4-ethoxy 4-amino 4-dimethylamino 2-iodo 2-hydroxy 2-phenoxy 2-methoxy 2-acetyl 2-ethoxy 2-amino 2-carboxy
2.2
0.9 1.1 1.3 na na na 0.6 na na na na na na na na 3.8 na 1.0 0.6
2.9 3.0
2.7 3.9 2.7 1.5
4.1 4.0 2.6 1.9 1.7 1.3 3.3 1.5
2.7 2.8 3.3 2.3 2.6 2.4 1.7 2.8 3.2 2.7 4.1 3.1
1.4 1.3 1.4 1.7 1.6
2.8 0.6 na 2.5 2.8 0.5 na 3.2 1.3 0.3 3.0 1.2 0.9 2.7 4.0 na 0.1 na na na na 3.7 2.0
2.0 1.6 1.1 1.2 1.2
1.0 0.8 1.0 0.7 0.7 0.7 0.9 0.6 na 7.7 na 4.4 2.0 4.5 3.5 0.4 na
1.2
1.3 na na
2.2 1.2 1.0
0.9 1.2 0.9 0.9 0.7 0.9 0.6 0.6 na 0.3 5.0 na 6.5 2.4 4.8 3.6 0.1 na
1.1
2.0 2.7 1.2
1.8 2.3 1.5 1.5
1.2
1.1
1.1 1.1 1.0
0.8 0.5 2.9 na 4.2 6.2 3.3 3.6 0.4 na
1.3 1.9 1.6
1.4 na 2.1 2.8
3.2 3.3 0.7 na
2.2
1.7 1.5 1.0 na 1.4 3.6 1.8 2.5 1.2
na
1.0
1.3 1.2 1.1
0.7 0.5 1.0 0.6 0.7 0.3 1.6 na 2.9 1.7 2.8 2.5 0.5 na
"kd in s-l. See Table I for conditions. na = not assessed.
of desorption of the solute from the sorbent surface. The plate heights were converted to kd values by use of eq 12. The rate constants so computed are quite small. In fact they are, in some cases, smaller than values reported for affinity chromatographic system^.^ However, the absolute desorption rate constants differ from these calculated values because of the peak asymmetry. Nevertheless, the relative values are important. As shown in Table V, the kd values are dependent on both the solute and the eluent. In the companion paper,l'l a wide variety of different eluents were studied and these trends become even more apparent. It is not possible to mechanistically rationalize this observation unless the probe solute and the eluent are both involved in the rate-determining step. By analogy to a conventional solution-phase displacement reaction (SN2),an intermediate should be invoked in which both the eluent and test solute are simultaneously bound to the surface. To arrive a t a more detailed hypothesis it should be understood that the zirconia used in this work is a farily crystalline monoclinic material in which the bulk zirconium ions are heptacoordinate. Assuming that the coordination number of the zirconium ions persists at the surface then structures of the following type will exist:
000
0-Zr-0
o/ ' 0
where the solid lines to oxygen atoms indicate bonds to structural sites in the bulk solid. In pure water, that is, in the absence of any foreign Lewis bases, the unsatisfied coordination sites of zirconium will be occupied by water, protonated water or by hydroxyl ions. Since our previous worklo indicates that at high aqueous concentrations of fluoride a minimum of three fluoride ions can bind per surface zirconium, we feel that it is reasonable to postulate at least three fairly labile coordination sites on each surface zirconium ion. This leaves the zirconium(1V) site with a formal charge of +2.26 In the presence of a Lewis base buffer one or more of the coordination sites of the surface zirconium ions, depending on the solution concentration, strength, and denticity of the
Lewis base eluent, is occupied by the eluent. Lewis base solutes will also bind to one of the coordination sites. Since both the eluent and solute are involved in the transition state, each is able to alter the strength of coordination of the other to the metal. If the eluent Lewis base is never displaced, it can still play a role in the kinetics by influencing the binding of the probe solute to a second site. This mechanism requires further investigation. Note that we argue that the better the eluent Lewis base is as a Lewis base, the better it elutes the solute. There are two possible explanations for this: (a) The eluent and solute Lewis bases occupy the same site, therefore, the eluent Lewis base leaves when the solute Lewis base arrives and vice versa. (b) The solute Lewis base is bound only to the few sites left exposed after the eluent Lewis base attaches to the surface. It is clear that the proposed mechanism is consistent with the very strong dependence of the kd data on both the type of solute and eluent. Retention in Citrate Eluents as a Function of pH. The retention of a series of p-benzoic acids in citrate media was examined as a function of pH. This study complements the preceding study in that the buffer system was fixed and the pH was varied, whereas in the above study we used a series of buffers of low Lewis basicity and the pH was adjusted by changing the buffer pK,. The results are summarized in Table VI and in Figure 5. Note that in comparison to the data in the other eluents (see Table I) retention in the citrate eluents is very weak. This is the result of two factors. First, 0.2 M sodium chloride was added to all the citrate buffers to minimize changes in the ionic strength as the pH was systematically varied. Second, and much more importantly, citrate is a considerably stronger Lewis base than either acetate or any amino sulfonate. At least at the higher pHs (see Figure 5) retention in citrate buffers increases with the solute's pK,. At pH 6.85 (see Table VI), the slope of a plot of log k'vs pH is 0.74, which is similar to the slopes in both acetate and the amino sulfonate buffers. However, as the pH decreased, the slope of this plot decreased until retention became independent of the the pK,. This is in distinct contrast to the behavior shown in Figure 1, which shows that the slopes are rather independent of the pH. At pH 5.28 the slope in citrate media decreased to 0.47. The lack of dependence on pK, at the lowest pHs (pH < 4) is almost
860
ANALYTICAL CHEMISTRY, VOL. 64, NO. 6, APRIL 15, 1992
Table VI. pH Dependence of Capacity Factors and Desorption Rate Constants of Benzoic Acids on Zirconia in Citrate Buffers" PH
(I
derivative
2.63
3.13
3.63
3.95
4.26
4.76
p-nitro p-cyano p-formyl p-chloro p-bromo p-iodo benzoic acid p-acetamido p-ethyl p-methyl p-isopropyl p-hydroxy p-ethoxy p-amino p-dimethylamino p-nitro p-cyano p-formyl p-chloro p-bromo p-iodo benzoic acid p-acetamido p-ethyl p-methyl p-isopropyl p-hydroxy p-ethoxy p-amino p-dimethylamino
1.04 (0.13) 0.76 (0.14) 0.73 (0.14) 0.71 (0.19) 0.82 (0.18) 0.97 (0.17) 0.31 (0.36) 0.49 (0.18) 0.47 (0.24) 0.39 (0.26) 0.46 (0.14) 0.33 (0.26) 0.59 (0.15) 0.90 (0.13) 1.95 (0.08) 0.10 (0.39) 0.09 (0.34) 0.11 (0.41) 0.19 (0.33) 0.19 (0.32) 0.21 (0.33) 0.11 (0.39) 0.19 (0.29) 0.19 (0.31) 0.20 (0.31) 0.13 (0.30) 0.28 (0.22) 0.37 (0.19) 0.39 (0.18) 0.71 (0.16)
0.81 (0.15) 0.64 (0.16) 0.64 (0.17) 0.70 (0.21) 0.79 (0.18) 1.06 (0.10) 0.30 (0.37) 0.48 (0.19) 0.47 (0.19) 0.40 (0.27) 0.44 (0.18) 0.34 (0.30) 0.59 (0.14) 0.65 (0.17) 1.45 (0.16) 0.10 (0.39) 0.09 (0.35) 0.10 (0.40) 0.18 (0.32) 0.18 (0.36) 0.19 (0.34) 0.11 (0.44) 0.18 (0.30) 0.18 (0.34) 0.19 (0.35) 0.12 (0.32) 0.27 (0.22) 0.34 (0.22) 0.38 (0.17) 0.67 (0.17)
0.60 (0.17) 0.50 (0.17) 0.54 (0.17) 0.71 (0.19) 0.78 (0.17) 0.90 (0.15) 0.32 (0.30) 0.52 (0.16) 0.50 (0.22) 0.44 (0.22) 0.45 (0.17) 0.40 (0.21) 0.67 (0.16) 0.59 (0.16) 1.23 (0.28) 0.10 (0.45) 0.07 (0.31) 0.10 (0.45) 0.15 (0.43) 0.15 (0.47) 0.16 (0.44) 0.11 (0.51) 0.16 (0.41) 0.16 (0.46) 0.16 (0.44) 0.12 (0.39) 0.24 (0.31) 0.27 (0.29) 0.32 (0.24) 0.52 (0.20)
0.49 (0.21) 0.43 (0.21) 0.47 (0.20) 0.69 (0.21) 0.74 (0.19) 0.84 (0.17) 0.34 (0.29) 0.57 (0.17) 0.54 (0.21) 0.49 (0.21) 0.45 (0.18) 0.48 (0.19) 0.76 (0.18) 0.62 (0.16) 1.17 (0.24) 0.09 (0.68) 0.06 (0.27) 0.11 (0.80) 0.12 (0.30) 0.11 (0.51) 0.13 (0.56) 0.09 (0.49) 0.11 (0.44) 0.12 (0.58) 0.11 (0.49) 0.09 (0.37) 0.17 (0.44) 0.19 (0.41) 0.20 (na) 0.32 (0.25)
0.29 (0.28) 0.26 (0.29) 0.30 (0.28) 0.50 (0.25) 0.53 (0.21) 0.62 (0.13) 0.28 (0.33) 0.46 (0.18) 0.45 (0.22) 0.43 (0.22) 0.36 (0.19) 0.48 (0.18) 0.76 (0.15) 0.58 (0.16) 1.07 (0.23) 0.01 (na) -0.01 (na) -0.01 (na) 0.06 (0.28) 0.08 (0.42) 0.07 (0.37) 0.01 (na) 0.08(0.38) 0.08 (0.42) 0.08 (0.43) 0.05 (0.23) 0.13 (0.50) 0.12 (0.43) na (na) 0.24 (0.32)
0.20 (0.45) 0.18 (0.42) 0.19 (0.40) 0.33 (0.30) 0.34 (0.27) 0.37 (0.18) 0.19 (0.40) 0.33 (0.23) 0.33 (0.26) 0.32 (0.25) 0.23 (0.26) 0.43 (0.20) 0.62 (0.18) 0.52 (0.17) 0.90 (0.16) -0.03 (na) -0.03 (na) -0.02 (na) 0.01 (na) -0.01 (na) 0.00 (na) -0.02 (na) 0.00 (na) -0.01 (na) 0.00 (na) -0.02 (na) 0.08 (0.37) 0.07 (0.31) na (na) 0.13 (0.34)
Values in parentheses are k d values in s-l. See Experimental Section for details.
1
1
0
001 0
c
-1
5
1
pH 2.63
o
0
o
0
I
J
1
/
-2.0
-2 5
1
1
35
I
I
I
43
45
50
I
pKo
Flguro 5. Correlation of the capacity factors for p-benzoic acids on rirconlum oxlde wilh pK.. All eluents contained 0.2 M sodium chloride and 20 mM citric acM at the indicated pH. The flow rate was 0.5 mLlmln at 35 OC. Slopes: (a) at pH 2.63, -0.01, sd 0.13; (b) at pH 5.28, 0.47, sd 0.07; (c) at pH 6.85, 0.74, sd 0.21.
certainly due to the fact that at these pHs, all of the benzoic acids are fully protonated. In essence, the proton has bested the surface zirconium site for occupying the valence of the Lewis base (carboxylate) site. Consequently, the Lewis basicity of the site, which is what induces the correlation with
pK,, is no longer of any importance. Thus, especially in view of the high ionic strength (0.2 M), it is unlikely that they are retained either by anion-exchange or ligand-exchange interactions. We conclude that in citrate medii, at the lowest pHs, retention is most likely due to hydrophobic and hydrogenbond interactions. The relatively high retentions of p-nitro- and p-cyanobenzoic acid (see Table VI) in citrate buffer show that interactions other than ligand exchange are partly responsible for retention. We attribute the difference in behavior in citrate buffers to the fact that citric acid is polyprotic and that ita state of ionization varies over the entire pH range. At low pH we presume that the surface Lewis acid sites are essentially fully populated with citrate or citric acid. Previous studies have shown that Lewis bases such as phosphate2' and fluoride1°J6 adsorb more strongly on zirconia at low pH. Closer examination of the data for citrate buffers reveals some interesting effects. Figure 6 shows the trends in capacity factor and desorption rate constants as a function of pH for p-cyanobenzoic acid, benzoic acid, and p-(dimethylamino)benzoic acid. Despite the complex trends, the values obtained are very reproducible and are very precise. As described above, all solutes show a consistent decrease in capacity factor as the pH is raised. Note that although these curves do not have maxima or minima, they show distinct inflection points and thus the dependencies on pH are not simple. In contrast, the desorption rate constants do not vary monotonically with pH. AU three derivatives have very high desorption rate constants at low pH. We believe that this is due to the complexity of the secondary interactions with the bound ligands. The rate constant then decreases as the pH is raised until the pH is approximately equal to the salute's pK,. A t this point, the desorption rate constants go through a maximum. This complex pattem in h vs pH is observed for all the benzoic acids tested here. We have no explanation for this phenomenon.
ANALYTICAL CHEMISTRY, VOL. 64, NO. 8, APRIL 15, 1992 2 5 ,
I
I
1
100 ' I
I
I
8
9
881
2 0 L
10 L
15
0
c
U LL
1 05
A
.-c 0 a 0
1
0
0
01
00 0.5
Zr02 I
3
I
I
I
I
4
5
6
7
0.01
5
6
7
PH
10
DH
10 L
0
+ U
LL 0
u
.
c
0
fY
r L
-1
6
3
0' 3
I
I
I
4
5
6
7
PH
Flgun 8. Plot of (a,top)log k'and (b, bottom) dewptkm rate constant for p-benzolc acids vs pH in citrate buffers on zirconium oxlde: (V) p~dlmethyIamln0)benzolc a W (0)benzoic acid; (0)pcyanobenzoic acid. Eluents contained 0.2 M sodium chloride and 20 mM citric acid at the appropriate pH. The flow rate was 0.5 mL/min at 35 'C.
However, previously groups have noted that the saturation capacity for the adsorption of carboxylic acids on metal oxide surfam goes through a maximum when the pH is nearly equal to the pKa.ss6 It is possible that these two phenomena are related. Comparieon to Bonded-Phase Ion Exchange. For comparative purposes, the benzoic acids were examined on a conventional silica-based strong anion exchanger. The results are given in Table III. The data clearly show that many of the interesting trends observed on the zirconium oxide phase are absent on the bonded-phase anion exchanger. There is no clear relationship between retention and pH on the silica-based exchanger. However, we could not explore as wide a range in pH on the silica-bad material as we did on zirconia due to the instability of the underlying silica packing material. In contrast to its behavior on zirconia, acetate buffer is actually a much weaker eluent on the bonded phase than is any amino sulfonate. In addition, the ortho isomeric benzoates were all easily eluted. This indicates that the chelation effect invoked above is not present on the bonded-phase material. Dicarboxylic acids were more retained than monocarboxylates; however, they are dianions and we therefore expect them to be more retained solely on the basis of electrostatic considerations. Thus many of the chemically interesting characteristics of the chromatography of anions on zirconia, which led us to propose the important role of surface Lewis acidity and ligand-exchange chemistry, are conspicuous by their absence on silica-based bonded-phase anion exchangers. There are other significant differences in anion chromatography on zirconia and this bonded-phase material. For example, in comparison to their behavior on zirconia, hydrophobic benzoic acids such as 1-naphthoic acid and 2-
0.01
I
SAX
5
6
7
I
I
a
9
I 10
PH
Flguro 7. Plot of log k' for aminobenzoic acids vs pH: (0)o-
aminobenzoic acid; (0)m-aminobenzoic acid; (V)paminobenzolc acid; (V)benzoic acid. Eluents were all amino sulfonic acid buffers (100 mM) with flow rates of 1.0 mL/mln at 35 O C . Regressions are least-squares fits. M),= zirconium oxide column; SAX = Spherisorb strong anlon exchange column. phenoxybenzoic acid are more retained on the bonded phase relative to leas hydrophobic solutes. This is undoubtedly due to interactions with the organic portions of the anion-exchange matrix. We also point out that on the silica-based supports at all pHs, all benzoic acids are easily eluted. In contrast, on zirconia at pH 6.1, the weakest eluent, we could not observe the elution of the more strongly retained solutes ( k ' > 100). Perhaps the most interesting point of comparison between the zirconia and the bonded-phase exchanger is the lack of any correlation between the solute's capacity factors and its pK, on the strong anion exchanger. This is shown in Figure 2 for data acquired at pH 6.8 and in Figure 3 for data acquired at pH 4.8. Both data sets include retention data for all isomers which eluted. The trend in log k'with pKa for the zirconium oxide phase is clear, but is totally absent for the strong anion exchanger. At pH 4.8, the capacity factors of the solutes on the conventional anion exchanger are larger, but there remains little overall selectivity related to the solute's pKa. The zirconium oxide data show the same trend as at pH 6.8. In general, the chromatographic selectivity for the benzoic acids is much better on the zirconium oxide phase than on the conventional anion exchanger. This is not surprising since the ionic properties of the solutes do not differ significantly while the geometry and Lewis basicity of the solutes differ greatly. The degree of isomer selectivity is shown more clearly in Figure 7. This shows the elution properties of benzoic acid and ita monoamine derivatives as a function of pH for both the zirconium oxide and the strong anion-exchange phases. The selectivity on the conventional anion-exchange phase is
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ANALYTICAL CHEMISTRY, VOL. 64, NO. 8, APRIL 15, 1992
low and varies erratically at the pH is changed. These nonsystematic variations in selectivity with pH may be the unfortunate result of the alkaline decomposition of the bonded phase under even these relatively mild conditions. The zirconium oxide phase shows great differences in retention behavior for the isomers. The o-amino derivative is capable of forming a chelate with Lewis acid sites and therefore shows the highest retention of any of the derivatives. The para isomer is able to interact via either the carboxylic acid group or the amino group and is relatively well retained. The meta derivative may be able to hydrogen bond to the surface while one functional group is coordinated to the Lewis acid site and is fairly well retained. Benzoic acid lacks any other functional group and is the least retained. This shows that the zirconium oxide phase is able to express more isomeric selectivity than does a conventional ion exchanger due to its ligand-exchangeinteractions as well as its hydrogen-bonding ability.
CONCLUSIONS The chromatographic properties of zirconium oxide toward benzoic acid derivatives differ substantially from those observed for a conventional silica-based strong anion exchanger. Chromatography on the zirconium oxide phase clearly involves a significant ligand-exchange contribution to retention which results from very strong Lewis acid-base interactions between the ionized carboxylate group serving as an electron-pair donor and surface ionic zirconium sites serving as an electron-pair acceptor. Selectivity among the various isomers is, in general, far greater with the zirconium oxide support than with the strong anion exchanger. This is demonstrated most dramatically by the differences in selectivity between 0-, m-,and p-hydroxy- and -aminobenzoic acid isomers, where both functional g-roups could interact with the ligand-exchange sites. Very strong eluents are necessary to elute the ortho isomers on the zirconium oxide phase while retention is very low on the conventional anion exchanger. The model of ion- and ligand-exchange interactions proposed by Regazzoni7to describe the electrophoretic characteristics of transition-metal oxides is readily adapted to qualitatively understand the present system once the Lewis base properties of the eluent are included. Ion exchange of noncoordinating solutes takes place a t ionized surface coordinated hydroxyl groups and water molecules. These equilibria also account for the decrease in ligand-exchangecapacity as the buffer pH increases.
ACKNOWLEDGMENT We thank Dr. Eric Funkenbusch and the Ceramic Technology Center at 3M for providing the zirconia particles used in these studies. We acknowledge funding from the National Science Foundation and the National Institutes of Health. J.A.B. acknowledges financial support from the Leading Edge Academic Program at 3M. Registry No. ZrO,, 1314-23-4; benzoic acid, 65-85-0.
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RECEIVED for review August 26,1991. Accepted January 21, 1992.
