Precolumn-Capillary Column Gas Chromatography for High Resolution Trace Determinations in Crude Extracts Wolfgang J. Kirsten, Per
E. Mattsson, and Harald Alfons
Department of Chemistry, Agricultural College of Sweden and National Swedish Laboratory for Agricultural Chemistry, S-750 07 Uppsala 7, Sweden
The sample Is InJected Into a packed precolumn and the solvent and low-bolllng compounds are eluted. The gas flow Is then dlverted Into a glass capillary trap, whlch Is cooled. After trapplng the analyte, the dlrectlon of the gas flow In the precolumn Is changed, the cooling Is removed, and the analyte Is eluted through the glass caplllary column. At the same tlme, the precolumn Is backflushed. When the chromatogram Is ready, the next sample can be Injected. Large volumes of crude extracts can be InJectedand traces of analytes be recovered from them quantltatlvely.
A precolumn-capillary column system with an internal cooling trap for the transfer between the columns was described by Kirsten and Mattsson ( I ) . Another system was proposed by Schomburg and Weeke (2). Both these systems were designed for metal capillary traps and columns, in which many classes of substances are catalytically decomposed already at low temperatures, and, in both systems, a quantitative transfer of the analytes to the capillary column is difficult because of the differences in diameter and flow rate between the packed and the open tube columns. We have further pursued the idea, perfected technical details, and applied the system to glass capillary traps and columns.
EXPERIMENTAL Apparatus. Flow diagrams of the system are shown in Figure 1. With the stopcocks in the injection configuration, the carrier gas passes into the apparatus through stopcock I and needle valve Q l , through cooling trap G, injection port S and precolumn A into detector D1 and out through stopcock 111. After the injection of the sample, this configuration is maintained until the analytes approach D1. Stopcock 111 is then turned to the trapping position and trap B is cooled. The gas now passes through the trap, where the analytes are condensed, via T-tube C out through stopcock 111. After trapping the analytes, the stopcocks are switched to the running/standby configuration, and the cooling is removed from the trap. Coming from stopcock I and cooling trap G, one part of the gas passes now through trap B and takes the analytes with it through the capillary column into detector D2. The other part passes backward through the precolumn, out through stopcock I1 and needle valve Q2 and sweeps out high-boiling fractions of the sample. When the chromatogram is ready, the stopcocks can immediately be switched to the injection configuration, and the next sample can be injected. A schematic layout of the arrangement is shown in Figure 2, and details are given in Figure 3. A Varian 2700 gas chromatograph with two Sc3H electron capture detectors is used. The exit opening of detector D1 is threaded and provided with a Teflon washer to give a gas tight connection to stopcock 111. There should be no resistance to the gas flow from the precolumn through detector D1 and stopcock 111. The fitting between the left inlet pressure regulator of the gas chromatograph and its injection port is opened and the regulator is connected to the Na-inlet of stopcock 1. The gas flow to stopcock I is thus controlled with this regulator. The line from stopcock I via valve Q1 and cooling trap G is connected to the injection port. The make-up gas NZto detector D2 comes from the instrument’s carrier gas inlet. Before entering into the system, the nitrogen from the tank is freed from oxygen by an activated copper 1974
catalyst. All stopcocks and needle valves are mounted on an extension of the front panel on the left side of the apparatus, and the cold trap G is mounted behind this extension. The injection port is provided with a septum spacer into which a hole is drilled into which a steel tube is welded for the connection to stopcock 11. The capillary trap B and the capillary column are connected to the precolumn resp. the T-tube (C) with shrink Teflon connections of the type shown in Figure 3 (HI.The tube of cooler, F1 or F2, can be inserted through a hole in the front of the apparatus, so that a part of trap B is inside it and is cooled by the carbon dioxide-air or the liquid nitrogen-air current. The hole is closed with lid E. Adjustment and Use of Apparatus. Turn on the gas flow and turn the stopcocks as shown in Figure 1. This configuration of stopcocks, called the running configuration, is also used for standby. Turn on the heat. Adjust the gas flow rates with the instrument regulator and with needle valve Q2 so that a suitable rate passes through the capillary column and a rate of at least 20 ml/min backwards through the precolumn. Open the way out from T-tube C through stopcock 111, close stopcock I1 and turn stopcock I so that the gas passes forward through the precolumn. Adjust the flow rate to about 10 ml/min with valve Q1. Open the way out from detector D1 through stopcock 111. The stopcocks are now in the injection configuration. Measure the flow rate, inject a clean test sample, and record the chromatogram with detector D1. Turn the stopcocks to the running Configuration. Read from the chromatogram the time interval during which the analytes are eluted. If the flow rate is much different from 10 ml/min, calculate the length of the interval which corresponds to this flow rate. Turn the stopcocks to the injection configuration and again inject a test sample. Record with detector D1. When the analytes approach, open lid E and insert cooler F. Open T-tube connection C through stopcock I11 and collect the substances in trap B during the calculated time interval. Turn the stopcocks to the running configuration, switch the recorder to detector D2, remove quickly cooler F and immediately close lid E. Record the chromatogram. Check that the total volume of gas which passes backwards through the precolumn in the running configuration per analysis is larger than that which passes through it in the injection configuration. If it is not, increase the opening of needle valve Q2 and reset the pressure of the instrument regulator, which has gone down, to its earlier value. The apparatus is now ready for the analysis of crude extracts, which is done in the same manner as described for the last test sample. If ghost peaks are obtained in blank runs-which is frequently caused by impurities in the carrier gas (1)--fill trap G with Dry Ice and acetone and keep it filled. Usually such ghost peaks cause no trouble, because they appear immediately after the removal of the cooler from trap B near the start of the Chromatogram. Overnight lower the temperature of the column oven to avoid deterioration of the capillary column. Over week-ends sweep out trap G. Turn stopcock I1 so that gas from the precolumn passes out directly to free air, and stopcock I11 so that the way out from detector D1 is open. Turn stopcock I so that the gas passes both ways, and remove the Dewar vessel from trap G.
RESULTS Figure 4 shows a chromatogram of a mixture of chlorinated hydrocarbons run through the described procedure. The separation corresponds to somewhat between 110,000 and 120,000 theoretical plates. Figure 5 shows a chromatogram of the hexane phase of a crude acetone-hexane-water extract of fish run with the above procedure. With a conventional method for pesticide
ANALYTICAL CHEMISTRY, VOL. 47, NO. 12, OCTOBER 1975
R
’
12 ....
=2
T A
Running /Standby
Figure 1. Flow diagram of
gas chromatographic system
(A) Precolumn. (6) Capillary cooling trap. (C) Capillary T-tube. (Dl.D2)Detectors. (G)Cooling trap for gas purification. (J) Capillary column. (N2) Carrier gas, nitrogen. (0) Needle valves for regulation of gas flow rates. (S)injection port with septum spacer
Q P2
Figure 2.
! .
Layout of apparatus
(A) Precolumn, i.d. 1 mm. length of filling 70 mm, exchangeable glass liner in injection port. (6) Capillary trap. (C) T-tube which is connected with silicone rubber tubing to stopcock lli and with a shrink Teflon connection to trap B and to capillary column J. (Di. D2) Electron capture detectors. (E)Lid for closing opening in front of oven through which the cooler (F1 or F2) is inserted. The lid is handled with handle (K).(G) Cooling trap for carrier gas. (J) Capillary column, Pyrex, 50 m. i.d. 0.3 mm, coated with SF96.(Ql, 03) Restriction needle valves. (I, 11, Ill)Stopcocks
analysis, traces of DDT in the order of magnitude of the detection limit-0.01 ppm-could just be discerned. Figure 6 shows chromatograms of sewage sludge extract run with a conventional packed glass pesticide column and with the described method. Well measurable sharp peaks are obtained with the capillary system whereas the conventional method only faintly indicates that something emerges from the column.
DISCUSSION A few years ago we had to determine extremely small residues of the herbicide picloram in forest soil. A number of peaks of unknown substances interfered with the gas chromatographic peak of picloram. Since the amounts of the interferences were too small for chemical or mass spectrometrical identification, there appeared to be only two solutions for this problem: Either to experiment haphazardly with tedious cleanup procedures, or to sharpen the chromatographic separation drastically. The second alternative was tried, because it appeared to give a basic solution also to other problems of this type. The only possibility to sharpen the separation sufficiently was obviously to use capillary columns. However, nonvolatile and high-boiling compounds, particularly polar high-boiling compounds, can spoil the separation and the column ( 3 , 4 ) .It appeared, therefore, necessary to avoid the introduction of such compounds with a precolumn. In our
Flgure 3. Details of
apparatus
(A) Precolumn with glass liner (R). (B) Capillary trap, two coils, inner diameter 0.3 mm, wall thickness 0.16mm. Thin walls are important for the rapid volatilization of trapped compounds. The trap is coated in the same manner as the Capillary column. (C)Low volume T-tube: A hole is pressed with a tung. sten wire into a heated glass capillary, i.d. 0.3, 0.d. 6 mm. The flat end of a piece of the same capillary is ground cylindrically to fit onto the wall of the other piece. The pieces are then sintered together as shown in the figure. (G) Dewar flask trap: Three coils of stainless steel tubing, i.d. 1, 0.d. 2 mm for each carrier gas line from stopcock I pass through the 1 I. Dewar flask with Dry Ice-acetone mixture. Dry Ice is added every morning and evening so that the trap is held cold continuously during a week. (E) Capillary connection to T-tube and precolumn, respectively. The opening of the thick-walled capillary end, which has been drawn out to an 0.d. of 3 and an i.d. of 0.3 mm is heated and widened with a tungsten wire, so that the capillary fits snugly into it with a negligible dead volume. The ends are then sealed together with shrink Teflon. (Fl)Dry Ice cooler for high-boiling analytes, (F2)Liquid nitrogen cooler for lower boiling analytes. Both coolers are Dewar containers with silvercoated vacuum insulation. A paper tube (M). which is prepared by rolling a paper sheet on a glass rod and glueing it, Is held concentrically inside the glass tube (L) with a few glued-on chips of a match. The flask (Fl) is filled with well-crushed Dry Ice and stoppered with cork stopper (P), and, when used, dry air is blown through it at a rate of 1.5 I./min. The cooler (F2)is charged with liquid nitrogen through joint (87) and dry air is blown through it at the same rate as used for F1. (N) Glass wool
first method ( I ) , we tried to solve this problem by injecting the sample extract into a packed precolumn outside the gas chromatograph and then trapping the analyte in a cooled zone a t the entrance of the steel capillary column. This system was rather complicated and many substances were decomposed on the metal surfaces. We tried then to mount both the precolumn and the capillary column into the same oven of the gas chromatograph and to transfer the sample from the precolumn into the capillary column with microvalves similar to those supplied by Carle (Carle Instruments, Inc., 1141 East Ash Ave., Fullerton, Calif. 92631), but made of gold and Teflon instead of glass and Teflon. DDT was used as the test substance. We had the same experience as Deans (5) that much of the sample disappeared in the Teflon of the valves. Such losses are probably dependent upon the nature of the analytes, since the successful use of such valves has been reported by several authors (6, 7). The new system was therefore designed so that there is no contact with the sample and other materials than glass, except, of course, the stationary phase. The capillary trap (B) was first made as a single coil. In order to find out if the analytes are quantitatively retained, the capillary column and the T-tube were removed and the trap was directly connected to detector D2. A solution of chlorinated hydrocarbons in hexane was then injected and run through detector D2. Temperature programming was used to obtain a sufficient separation. The same chromatogram was then run again with the carbon dioxide cooler
ANALYTICAL CHEMISTRY, VOL. 47, NO. 12, OCTOBER 1975
1975
cool.;
tI rnappc tcold lo"'
switched tiow
Oft
Figure 4. Mixture of commercial chlorinated hydrocarbons run through the procedure. Precolumn: 5% SF96 on Chromosorb G, AW/DMCS, 80-100 mesh. Cooling trap one coil. Capillary column 50-m Pyrex. i.d. 0.3 mm coated with SF96. Flow rates: Precolumn forward 6 mi/min, backward 20 ml/min, capillary column 1.7 ml/min, make-up gas in detector (D2) 20 mi/min. Temperatures: Oven 185 'C, injector 205 OC, detectors 205 OC. Sample: 0.4 pl of hexane solution containing 19 pg of lindane, 26 pg of heptachlor, 44 pg of aldrin, 48 pg of p,pDDE, 57 pg of o,pDDD, 140 pg of p,p-DDD, 32 pg of p,pDDT.
i
cooler O f f
start t l o w switched
50 m i n
LO mtn
30 min
20 m m
Flgure 5. Crude fish extract run through procedure Conditions as in Figure 4. Sample 10 MIof hexane phase of acetone-hexane-water extract
1976
ANALYTICAL CHEMISTRY, VOL. 47, NO. 12, OCTOBER 1975
10 min
t r a p cold
-
I-
Y,"
53
LO
30
20
3
1
Figure 8. Chromatograms of sewage sludge extract run with a packed column and with the described method. Sample prepared for determination of chlorlnated hydrocarbons by extraction with hexane-acetone-sodium chloride in water solutlon, concentration of the hexane phase and treatment with fuming sulfuric acid. The hexane phase was then treated with a drop of mercury to remove sulfur. Chromatographic conditions: Packed column length 3 m. i.d. 1.5 mm, 5% OV101 -k 7.5% QF1 (1 3) both on Chromosorb W, AW HMDS. Column, Injector, and detector temperatures 185, 230 and 215'C, respectively. Nitrogen flow 20 ml/ min. Sample 1.3 gl, Conditions for capillary column system as in figure 4, except that the capillary gas flow was 2 ml/min. Sample 3 gl. Retention times for DDT and its metabolites and lindane are indicated with numbers 1-6. Most of the other sharp peaks of the chromatograms correspond to different polychlorinated biphenyls
+
(F1) inserted. The curves are shown in Figure 7. There is no indication of any peak in the second chromatogram before the cooler is removed, which shows that there is no break-through of the analyte. There might, however, be a slow bleeding. In order to check this, a mixture of chlorinated hydrocarbons was run several times under the same conditions, except that the time of cold-trapping was varied in the order 10,5,20, and 5 minutes. If there was bleeding, one would expect that the runs with longer-time trapping should give lower yields than those with shorter-time trapping. One would also expect that the ratios between earlier peaks and later peaks in the same run would change with the time of trapping, because the bleeding would
20 m,n
15 rn,"
probably affect the lower boiling substances more than the higher boiling substances. Twenty microliters of the sample solution were injected during 20 seconds every time. A high chart speed, 50 mm/ min, was used, and the peak areas were determined by the cut-and-weight method. The peak areas obtained are reported in Table I and the ratios between the peaks in Table 11. There is no indication that the runs with longer trapping times give lower results, nor is there any indication of changes of the peak ratios. It appears, therefore, probable that there is no bleeding from the trap under the conditions used for these measurements. Further experiments showed that the solvent, hexane, is not retained in the trap with carbon dioxide, nor is 1,1,2,2tetrachloroethane, bp, 146.3 "C. Lindane boils with decomposition at 288 "C. In order to be able to determine lower boiling substances, the liquid nitrogen cooler (F2) was made. 1,1,2,2-Tetrachloroethane did not break through the trap which can be seen in Figure 8A. When we tried to find a somewhat higher boiling compound for these experiments, we found 2,4,6-collidine on our shelf. Since the electron capture detector has a low sensitivity for this compound, we had to use much more of it. The results are shown in Figure 8B. Obviously, collidine broke through the trap. However, when the substance ceased to come from the precolumn, nothing more came out from the trap. Hence, the break-through was not caused by a too high vapor pressure of collidine, but rather by the formation of an aerosol. When a molecule of the analyte moves in the capillary and is suddenly cooled down, there are two chances: Either it hits the wall with the stationary phase and is held back, or it hits other molecules and forms clusters or crystals which are not held back but pass through as an aerosol. Obviously, the chance to hit the wall first must be much greater if the concentration of the substance is low. This is probably the main reason why the chlorinated hydrocarbons were quantitatively retained and collidine was not. In order to obtain a better retention of larger amounts of
10 min
5 mtn
Figure 7. Retention test of cooling trap. Mixture of commercial chlorinated hydrocarbons run through precolumn and trap only. Precoiumn: 5 % SF 96 on Chromosorb C, AW/DMCS, 80-100 mesh. Injector, 205 OC; detector, 205 OC; nitrogen flow, 6 ml/min. Sample 0.4 Mi of hexane solution, single coil trap, Dry Ice cooling, temperature programming. Start 140 OC, 8°C/min, rnax. 195 OC.One curve with cooled trap, one without cooling
+
ANALYTICAL CHEMISTRY, VOL. 47, NO. 12, OCTOBER 1975
1977
A
B
N
1
J j ; 5 rnin
5 min
lo'min
Y
8.
u o
5 min
Figure 8. Retention tests of cooling trap. Liquid nitrogen cooling. Gas flow rate in column and trap: 6 ml/min. Temperatures: injector 190 OC. detector 190 O C , oven 35 OC. Samples: (A) 0.2 fit of hexane solution containing 20 ng of 1,1,2,2-tetrachloroethane.(B) and (C) 2 fil of hexane solution containing 6 k g of 2,4,6-collidine. Curves A and B single coil trap: tetrachloroethane is quantitatively retained, collidine breaks through. Curve C double coil trap: collidine is almost quantitatively retained
Table I. P e a k Areas Obtained in Retention Tests of Cooling Trap min
Peak l a
Peak 2
Peak 3
1 10 1.8 15.6 12.7 2 5 2.1 17.7 15.5 3 20 1.9 15.6 13.8 4 5 1.6 13.7 11.9 a Peak 1 is heptachlor, retention time 10 minutes; peak 2 aldrin, 13 minutes; and peak 3 is an incompletely separated double-peak of p,p-DDE and o,p-DDD with retention times of 25 and 25.5 minutes, respectively.
analyte, trap B was redesigned as a double coil trap. The amount of analyte which passes through the first coil is small. When passing out from the first coil it is vaporized, and then it passes into the second coil with a low concentration, and, because of the reasons given above, a better retention is obtained, which is shown in Figure 8C. Obviously a negligible trace also passes through the second coil. These experiences with cooling traps agree well with those reported by Buchanan and Nakao (8) for large cooling traps in elementary analysis. Since the amount of collidine used in these analyses-6 pg-was enormous compared with the picogram quantities which are usually determined, and which were quantitatively retained already in one coil, we did not introduce a third coil. The question arises, however, what happens if a small amount of analyte passes into the trap along with large amounts of other compounds? I t appears probable that a part of the analyte will be caught on aerosol particles of the other compounds. In this case, it might be necessary to use more coils. The use of more than one coil for the trapping does not 1978
Ratios, peak areaipeak area Cooling time,
Peak areas, c m 2
Cooling time,
Rm KO,
Table 11. Ratios between P e a k Areas Reported in Table I Run KO.
min
1/2
113
1 2 3 4
10 5 20 5
0.12 0.12 0.12 0.12
0.14 0.14 0.14 0.13
213
1.2 1.1 1.1 1.2
impair the peaks of small amounts of analytes, which are already retained in the first coil. It must, however, cause a broadening of peaks when parts of analytes are first retained in the second or later coils. The distance of the condensation zones of the two coils in our apparatus is about 20 cm. The length of the capillary column is 50 m. The peak broadening caused by one additional peak can, therefore, be calculated to be about 0.4% of the retention time. If we apply this to Figure 4,it means that the width of a lindane peak would increase with 0.1 mm and that of a p , p DDT peak with 0.4 mm. The broadening would, however, impair only peaks which are much larger than those of the figure. Though this broadening appears tolerable in this case, it might become objectionable in cases when very impure extracts are analyzed, and when small amounts of analyte pass into the trap along with other compounds and are retained quantitatively only in two or maybe more coils. If an uncoated capillary trap were used, most analytes would probably not migrate through the trap, but they would be carried through it with the full velocity of the gas flow. If we again assume the conditions of Figure 4, this would mean that the 20 cm of the trap would be passed in 0.5 second, and the resulting peak broadening for every additional coil would be 0.02 mm regardless of the retention time. The peak broadening would thus be negligible even if several coils were used.
ANALYTICAL CHEMISTRY, VOL. 47, NO. 12, OCTOBER 1975
In the earlier stages of this work, we used uncoated capillary traps with good results. Because of theoretical considerations-we supposed that the coating would give a safer retention and that it would diminish the risk of decomposition of the sample-we changed to coated traps, and all the reported work was done with coated traps. It appears, however, that uncoated traps might be preferable in many instances. (At the time of printing, we had made experiments with a three-coiled uncoated trap which showed that a t least 320 pg of a mixture of different tetrachloroethanes can be trapped quantitatively. No experiments have yet been made to evaluate the peak broadening.) The precolumn was expected to become contaminated and spoiled very soon. I t was therefore designed to be simple and easily exchangeable. However, after three months' use, clean chromatograms were still obtained, and no serious interferences from contaminations were stated, except that the DDT peaks from standard test solutions became somewhat smaller and the DDE peaks somewhat larger. This indicated decomposition of DDT in the injection port. Large deposits from extracts were found there when the column was taken out, but the column filling was still in good condition. The precolumn was therefore redesigned to incorporate an exchangeable glass liner as shown in Figure 3, so that deposits easily can be removed without removal of the column. The described precolumn-capillary column system can obviously be modified in many ways. Right now we are assembling an arrangement in which the precolumn is not connected to detector D1 but via a T-tube and an an-off
topcock to detector D2. Since this line is used only to blow off the solvent and low boiling compounds and to determine the retention times of the analytes in the precolumn, losses of analyte can be tolerated there. The arrangement makes it possible to use an ordinary one-channel gas chromatograph and also to use other types of detectors. The handling of the equipment can undoubtedly be simplified considerably by the use of a multichannel stopcock instead of several two-way and three-way stopcocks, or by the use of automatic control equipment as described by Schomburg and Weeke (2).
ACKNOWLEDGMENT The authors are indebted to Nils Larsson for valuable glass blowing designs. LITERATURE CITED W. J. Kirsten and P. E. Mattsson, Anal. Lett., 4, 235 (1971). G. Schomburg and F. Weeke, "Gas Chromatography 1972", Applied Science Ltd., Barking, Essex, England, 1973, p 285. (3) K. Grob and J. J. Jaeggi, Chromtographia, 5, 382 (1972). (4) A. L. German and E. C. Horning, Anal. Lett., 5, 619 (1972). (5) D. R. Deans, Chromatographia, 1-2, 18 (1968). (6)E. C. Hornlng, C. D. Pfaffenberger, and A. C. Moffat, Anal. Chem., 44, 2
.- .
11973\ I _,.
(7) D. C. Fenimore, R . R . Freeman, and P. R. Loy, Anal. Chem., 45, 2331 (1973). ( 8 ) D. L. Buchanan and A. Nakao. J. Am. Chem. Soc., 74, 2389 (1952).
RECEIVEDfor review March 17, 1975. Accepted June 30, 1975. The work was sponsored by the Research Committee of the National Swedish Environment Protection Board.
Kinetic Measurements by Reactions during the Overlap of Elution Zones with Different Elution Speed in GasChromatographic Columns Paul Schulz Centrul de Chimie Timisoara, Romania
According to a classical method of kinetic measurement on liquid phase reactions by gas-chromatographic procedures, the lower volatility reaction partner Is previously dissolved in an adequate liquid phase. The volatile reaction partner is then Injected into a gas-chromatographic column containing this liquid phase. Using a somewhat different method, which may be called a "double-pulse technique", both reagents, which must have different elution characteristics and must react irreversibly, are injected consecutively into a gaschromatographic column containing an adequate liquid phase. The contact occurs by overlap of the elution zones in the column. An attempt is made to interpret the resulting conversion in the special case of second-order reactions. Kinetic data obtained with the esterification of mxyienoi with acetic anhydride are in fair agreement with values obtained by a classical method. Perspectives and limitations of this procedure are discussed.
Several techniques were described for catalytic and noncatalytic reactions in gas-chromatographic columns. Some procedures allow the investigation of liquid phase reactions
between a nonvolatile component dissolved in the liquid phase of the column and a volatile one injected and eluted through it. A number of irreversible reactions, e.g., the Diels-Alder reaction ( I , Z ) , the oxidation of trialkylphosphites (31, esterifications ( 4 ) , etc. were investigated using this technique. Because of the special requirements on the elution properties of the reagents involved (e.g., the magnitude of their reaction rate), this procedure is not widely applicable and is restricted to several special cases. It would be applicable for relatively fast reactions except for the limited relaxation time of mass transfer between phases in the gas-chromatographic column. These times are usually from to 1 second ( 5 ) .Under satisfactory conditions, reactions with half-times of about 10 seconds could be investigated without mass transfer perturbations ( I ) . A different technique to use for the reacting components to make contact in the chromatographic column is to inject the slowly eluting components first, and then after an adequate time, the components that elute faster. Contact and reaction occurs by the overlap of the eluting zones. Reaction should be fast enough to give measurable conversion, but should not approach diffusion control.
ANALYTICAL CHEMISTRY, VOL. 47, NO. 12, OCTOBER 1975
1979
