Anal. Chem. 1983, 55, 1188-1191
1188
Postcolumn Deoxygenator for Liquid Chromatography with Reductive Electrochemical Detection Robert E. Reim Analytical Laboratories, 1602 Building, The Dow Chemical Company, Midland, Michigan 48640
T o date applications of liquid chromatography with electrochemical detection (LCEC) have been primarily to the determination of trace organic species such as neurological agents, pharmaceuticals, and antioxidants via oxidation techniques (1). Unlike oxidative LCEC, relatively few applications of reductive LCEC have been reported even though a large number of compounds are amenable to reductive detection (2). One problem which has contributed to the slow growth of reductive LCEC is the need to exclude dissolved oxygen from the chromatographic system or discriminate against its effects. In acid medium, oxygen is easily reduced and the reduction proceeds in two steps Oz 2H+ 2e- HzOz E l l z = -0.05 V vs. SCE (1) HzOz 2Hf 2e2Hz0 Ellz -1.0 V vs. SCE (2) The presence of oxygen in the mobile phase, about 0.5 mM in air-saturated aqueous solution, therefore results in high background current. Also hydrogen peroxide, an intermediate in the reduction process, may chemically react with components in the mobile phase. Several methods of removing dissolved oxygen have been reported including rigorous purging of mobile phase and sample solution with inert gas ( 3 , 4 ) ,pre- or postcolumn addition of a chemical reductant (5), and preferential electrochemical reduction (6, 7). Methods of discriminating against the effects of dissolved oxygen have also been reported including reverse pulse amperometry (8) and dual electrode detection (9). This paper describes a novel method for the continuous removal of dissolved oxygen from chromatographic effluent, based on the use of a tubular semipermeable membrane.
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EXPERIMENTAL SECTION Apparatus. Data were collected with a flow injection system consisting of an Altex Model llOA pump, a Laboratory Data Control loop injector, a suitable membrane device, and an EG&G Princeton Applied Research (PARC) Model 310 static mercury drop electrode (SMDE) with associated PARC Model 174A potentiostat and a Linear Model 355 strip chart recorder. In preliminary studies the membrane device consisted of a 500-mL vacuum flask containing the tubular membrane of interest. The membrane was connected into the flow system via standard stainless steel chromatographic fittings which were mounted in a rubber stopper. Vacuum was supplied by a Sarvac Model 8804 vacuum pump. A McLeod gauge was used to monitor pressure within the flask. In later studies the deoxygenatingdevice shown in Figure 1 was used. The clear plastic shell of the device (3.81 cm X 4.13 cm X 10 cm) was fitted into polypropylene end caps (radius 5.08 cm) via a double O-ring seal. The assembly was held together with four threaded screws. One polypropylene cap contained a fitting for vacuum pump connection and a second fitting for bleeding the shell. The other cap contained standard male chromatographic fittings through which a tubular silicone membrane typically 0.25 mm X 0.63 mm X 3.0 m was sealed with silicone rubber sealant. Sealant was also used to fashion suitable flared connections on the ends of the membrane tube. Silicone elastomer tubing was supplied by Patter Products, Beaverton, MI. Chromatography. For chromatographicstudies in the cation exchange mode, the following conditions were used: column, 3
mm X 150 mm surface sulfonated styrene divinylbenzene X 2, 50 pm; injection volume, 150 pL; detector, PARC Model 310 SMDE; drop time, 0.5 s; drop size, large; detector potential, -0.8 V vs. Ag/AgCl; shell pressure, 1 mmHg. For chromatographic studies in the reversed-phase mode, the following conditions were used: column, Chromatix RP-18, 10 pm, 4.6 mm X 250 mm; eluent, 0.1 M NaH,P04-0.002 M H,PO, in 50/50 acetonitrile/ water (v/v); flow, 1.1mL/min; injection volume, 20 pL; detector, Bioanalytical Systems TL-6 mercury/gold amalgam, detector potential, -1.0 V vs. Ag/AgCl. RESULTS AND DISCUSSION The theory for ideal gas permeation in a static gas/membrane/gas system with fixed gas transport to the inner wall has been reviewed by Yasuda (IO). The gas flux across the membrane is
AP (3) 1 where P = permeability coefficient, cm3 cm s-l cm-2 cmHg, Ap = pu- p z where p1= pressure at inner membrane wall (cm) and p z = pressure a t outer membrane wall (cm), and 1 = membrane thickness (cm). For ideal systems, the permeability coefficient of the membrane to a dissolved gas is essentially the same as that of dry gas in the gas/membrane/gas situation. In practice, however, boundary layer resistance can lower the dissolved gas transfer rate through the membrane. With tubular membrane geometry the boundary condition used to derive eq 3 (i.e., fixed mass transport) no longer holds since the dissolved gas concentration and wall flux vary along the tube length. Mass transfer expressions have been derived (11, 12) for the case of stationary diffusion in laminar flow in a tube and the overall mass transport rate is reportedly determined by membrane resistance together with liquid phase resistance (12),Le., diffusion of the dissolved gas to the inner membrane wall. Preliminary Studies. Several tubular polymeric membranes were tested for their ability to permeate dissolved oxygen by using the vacuum flask technique described in the Experimental Section. The membranes tested were silicone rubber, Teflon, 4-methyl-l-pentene, Tygon, and Nafion. Membranes were chosen to be of high to low permeability, respectively, based on ideal gas permeability data (10). With the exception of Nafion all the membranes tested were nonporous. Because of its high oxygen permeability and chemical inertness relative to the other membranes tested, silicone rubber was chosen for further testing. Several approaches were considered to minimize the external membrane pressure since oxygen transport is directly proportional to the pressure difference across the membrane. Among the approaches considered were purging of the shell with inert gas, evacuation of the shell and countercurrent flushing of the shell with alkaline-sulfite solution. All of the approaches were effective for removing oxygen; however, evacuation was slightly superior in performance and proved to be more practical. Inert gas purging required large volumes of gas to rapidly flush the shell. Alkaline-sulfite solution, which potentially offered maximum Ap sinc pi = 0, had the lowest oxygen removal (92%) possibly because of slow reaction kinetics at the outer membrane wall. In addition the pulsing action of the countercurrent pump was transmitted to the detector, increasing base line peak-to-peak noise 10-fold.
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0003-2700/83/0355-1188$01.50/00 1983 Amerlcan Chemical Society
ANALYTICAL CHEMISTRY, VOL. 55, NO. 7,JUNE 1983
1189
Figure 1. Drawing of deoxygenator. Vacuum
Residence Time [seconds)
Figure 4. Dissolved oxygen concentration as a function of residence time: (A) silicone rubber, 0.30 mm X 0.63 mm X 3.0 m; (B) silicone rubber, 0.51 mm X 0.94 mm X 3.0 mm; (C) theoretical.
: E Time (minutesl
Figure 2. Removal of dissolved oxygen from 0.1 M HC10, with deoxygenator: membrane, 0.30 mm X 0.63 mm X 3.0m silicone; flow, 1.5 mL/min; shell pressure, 0.9 mmHg; detection potential, -0.5 V.
j L
T-iA 1
0
1 2
I 4
1 6
I I
1 8
1
1
1 0 0 2 Time (minutes)
l
I
4
l
6
I
8
I
1
0
Figure 5. Chromatogram using deoxygenator: (A) 1 ppm copper(I I), (B), air-saturated blank; mobile phase, 0.1 M HCIO,; flow, 1.2 mL/rnin; background current, 110 nA. Table I. Comparison of Dissolved Oxygen Concentration shell pressure, mmHg 222 181 139
0
05 Potential I V I
10
Figure 3. Current-voltage curve showing removal of dissolved oxygen from 0.1 M HCi04: (A) air-saturated; (B) 0.9 mmHg shell pressure.
Performance. The removal of dissolved oxygen from 0.1
M HCIOl with a silicone rubber membrane in the shell and tube design is shown iin Figures 2 and 3. At E,, = -0.50 V the large cathodic current due to reduction of dissolved oxygen rapidly decreases with evacuation of the shell and stable background current is achieved after about 25 min. The i-E curve, Figure 3, shows dissolved oxygen concentration is significantly reduced. In this example about 98% of the dissolved oxygen concentration h w been removed. The effects of flow rate and tubing geometry on dissolved oxygen removal at fixed pressure are shown in Figure 4. Silicone rubber tubes of different internal diameter and wall thickness but of same length were tested at varying flow rates. Oxygen concentration wa8 calculated after measuring the current due to &-saturated carrier solution and the current due to deoxygenated carrier solution at E = -4.5 V. Data were corrected for the flow rate dependence of the SMDE. For shorter tube lengths of a given diameter the fraction of oxygen removed was inversely pro-
98 56 44 31 15 4
ppm dissolved oxygen exptl theoretical 2.8 2.2
2.5 2.1
1.7
1.6 1.1
1.2 0.60
0.64
0.46
0.50
0.28 0.15
0.35 0.17 0.05
0.08
portional to flow rate. The concentration of dissolved oxygen is apparently fixed by the kinetics of mass transport to the inner membrane wall. With sufficient residence time the minimum concentration of dissolved oxygen was in agreement with that predicted from the thermodynamic limit of the system, Henry's law
C = kp
(4)
where C = concentration of dissolved oxygen (ppm), p = partial pressure of oxygen (cmHg), and k = constant, 0.54 a t 25 OC. As shown by additional experimental data in Table I, the effect of external membrane pressure, p z on dissolved oxygen concentration (current) is in general agreement with that calculated by using eq 4. Calculation, therefore indicates that for 99.9% oxygen removal in an ideal system an external membrane pressure of 0.18 mmHg is required. For the apparatus reported here, a minimum shell pressure of 0.5 mmHg was achieved. Chromatography. Typical examples of ion exchange separations obtained with the postcolumn deoxygenator are shown in Figures 5 and 6. Without removal of dissolved
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ANALYTICAL CHEMISTRY, VOL. 55, NO. 7, JUNE 1983
I*
Table 11. Removal of Dissolved Oxygen from Eluted Supporting Electrolytesa back% ground cathodic oxygen current, potential electrolyte removed nA limit, V 0.1 M HClO,
1 I
0.1 M KNO, 0.1 M NaOH 0.02M H,PO, in 50/50 methanol/ water (v/v) 0.02 M H,PO, in 65/35 acetonitrile/ water (v/v)
lOnA
I
99 99 98 94
110 90
87
980
-1.0
-1.4 -1.7
60 640
-1.2 -1.2
a Silicone rubber membrane, 0.30 mm X 0.63 mm m; flow, 1.5 mL/mia; detector potential, -0.8 V.
,!, 0
2
,
, ,
, , ,
4 6 8 10 Time (minutes)
12
X
3.0
I1
,
14 16
Flgure 6. Chromatogram using deoxygenator showing separation of cadmium and lead: (A) 1 ppm Cd(II), (E) 1 ppm Pb(I1); mobile phase, 0.1 M HCD4-0.005 M Mg(CIO4)&flow, 1.0 mumin; background current, 140 nA.
-
A 0
4
8
b
0
4 8 Time (minutes)
0
4
8
Flgure 8. Peak broadening effect of deoxygenator (void volume 205 pL), for ion exchange separations: (A) without deoxygenator; (E) with deoxygenator, 775 mmHg; (C) with deoxygenator, 0.5 mmHg.
0
"4
8 12 16 Time Iminutesl
20
24
Figure 7. Chromatogram using deoxygenator showing separatlon of nitroaromatic compounds: (A) 60 ng of 4-nitrophenol; (E) 20 ng of 2-nitrophenol; (C) 40 ng of nitrobenzene; (D) 60 ng of dinltro-o-secbutylphenol.
oxygen, quantitation of 1 ppm copper(I1) ion by reductive LCEC was impossible because of the high background current (13 pA a t E = -0.8 V) and large peak-to-peak base line noise (600 nA). With postcolumn oxygen removal, the copper peak was well-defied and sub-part-per-millionconcentrations were easily quantified. The background current was reduced to 110 nA and base line noise was reduced to
