Double-Column Programmed Temperature Gas Chromatog ra phy Volatile Polar Column Packings and Quantitative Aspects EDWARD M. EMERY and W . E. KOERNER Research Deparfmenf, Organic Chemicals Division, Monsanfo Chemical Co., St. Louis 77, Mo.
b The double-columr programmed temperature technique in gas chromatography has been extended to more polar and volatile column packings. Comparisons of single- and doublecolumn base-lines as well as the advantages of the double-column technique for extending the upper temperature limit on such column packings are shown. The effect of column substrate in the sample and reference gas streams due to bleeding at the high temperature end of programs with volatile column packings was studied. No effect on detector response was found.
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programmed temperature gas chromatography the vapor pressure of the substrate increases as the temperature of the column is raised and causes an upscale drift in the base-line which limits the maximum temperature to which many otherwise thermally stable column packings can be programmed. Previously (1) we described an instrument using the doublecolumn technique in which the carrier gas is split into two streams which are passed in parallel through a pair of symmetrically heated columns and then through the reference and sample sides of a symmetrical thermal conductivity detector. The vapor pressure increase of the column packing is compensated and a level base-line can be achieved up to much higher temperatures. In this paper two other aspects of double-column programmed temperature gas chromatography are considered. The extension of the technique to volatile polar packings, an especially attractive area of utilization, is demonstrated for two useful polar packings for analyses which are of wide potential interest in biological chemistry. Secondly, the problem of constancy of relative response in the presence of varying levels of vaporized liquid substrate was studied to ascertain whether special problems would be encountered with quantitative analyses in double-column programmed N SINGLE-COLUMN
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temperature gas chromatography. If the presence of column substrate in the sample and reference gas streams a t the high temperature end of a program had an appreciable effect on the detector response, then it would be very difficult to do quantitative work by such a technique since the response of the detector for components eluted when the columns are bleeding would vary with the heating rate, gas flow rate, and the extent of bleeding a t each temperature point on the program. INSTRUMENTS AND OPERATING CONDITIONS
Two early prototypes of the F&M Scientific Corp. Model 720 gas chromatograph were used in these studies. Both instruments were double-column programmed temperature gas chromatographs which utilized symmetrical hot-wire thermal conductivity detectors. A 1-mv. range Brown recorder equipped with a Model 201 Disc Integrator was used to record the chromatograms and integrate the peak areas. Operating conditions are summarized below : Detector temperature, 300" C. Bridge current, 150 milliamperes. Injection port temperature, 310" C. Sample column flow rate, 60 to 140 cc. per minute. Reference column flow rate, adjusted empirically to give good base-lines in blank runs. Column, 1 or 2 meters X 1/4 inch 0.d. stainless steel tubing. RESULTS AND DISCUSSION
Column Packings. The chromatograms shown in Figure 1 were obtained a t an attenuation of 1 with a pair of 2-meter columns packed with 20 weight % LAC-3R-728 polyester (Cambridge Industries Company, Inc.) on 60- to 80-mesh Siliconized Chromosorb W. A sample column flow rate of 140 cc. per minute was used. Base-line performance obtained under a single-column arrangement where the reference column is replaced by anempty tube is shown in Section A of Figure 1. The comparison shows that it would be difficult to do programmed temperaVolatile
ture work much above 200' C. on a single column of this volatile but highly selective packing. Section B of this same figure shows a chromatogram obtained under the same conditions from a 0.004 ml. portion of a 10% solution in ether of a known mixture of fatty acid methyl esters. The first figure in the subscript represents the number of carbon atoms and that foIlowing the colon represents the number of double bonds in the acid portion of the ester. The saturated acid esters list only the number of carbon atoms in the subscript. The chromatogram shows good resolution of the four Cia fatty acid methyl esters (stearate, oleate, linoleate, and linolenate) and elution of even the Cg4 saturated ester. Section C of this figure shows a chromatogram obtained under these same conditions from a 0.004-ml. portion of a 10% solution in ether of a mixture of methyl esters of cod liver oil fatty acids. It shows two components eluted beyond the retention time of the Cu saturated ester. When this mixture was chromatographed isothermally a t 185" C. on this same column packing, the various Cis acid esters were eluted in about this same time with comparable resolution, but the last component shown here a t 24.5 minutes took over an hour to be eluted and had such a long low peak shape that it would have been difficult to determine quantitatively. Yet this highly selective substrate needed for the c 1 8 ester resolution could not be programmed much above 200" C. by the usual single-column technique. All of the chromatograms shown in Figure 1 were obtained on freshly prepared packing. After longer use the doublecolumn base-line shows even greater stability while still giving good resolution of the (218 esters. Operation to these high temperatures with such a volatile substrate does, of course, shorten the life of the column packing. However, as the exposure to the high temperature is only brief during each analysis, the column life is not so short as might be expected. Under such use the columns will last for a t least 50 analyses.
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Figure 2. Double-column programmed temperature chromatograms on Tween 80-0-phosphoric acid column packing A. B.
Comparison of single- and double-column base-lines Known mixture of free fatty acids
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Figure 1 . Double-column programmed temperature chromatograms on LAC-3R-728 polyester column packing A. 6. C.
Comparison of single- and double-column base-lines Known mixture of fotty acid methyl esters Methyl esters of cod liver oil fatty acids
The chromatograms shown in Figure 2 were obtained at a n attenuation of 1 with a pair of 1 meter columns packed with 20 weight yo Tween 80 plus 2 weight % o-phosphoric acid (85%) on 60- to 80-mesh Chromosorb K. A sample column flow rate of 120 cc. per minute was used. Section A of Figure 2 shows a comparison of double- and single-column base-lines obtained on the Tween 80phosphoric acid packing. This very polar column packing cannot be programmed satisfactorily much above 150" C. under the usual single-column method. Section B of this figure shows a chromatogram obtained under the same conditions from a 0.006-ml. portion of a 10% ether solution of a known mixture of free fatty acids. The mixture contained all of the normal saturated free acids from C1 (formic) through CIa and also isobutyric and isovaleric acids. A 2-meter column of 20 weight yo LAC-3R-728 polyester and 2 weight yo o-phosphoric acid on 60- to 80-mesh Chromosorb W a t low temperatures
will not resolve these is0 acids from the preceding normal acids. Quantitative Response Studies. T o study the bleeding effect quantitatively, a binary sample mixture was chosen such t h a t on a given volatile column packing the first component was eluted early in the program when the packing was not bleeding appreciably and the second component was eluted at the high temperature end of the program when the packing was bleeding heavily. An equal weight mixture of naphthalene and p-terphenyl, dissolved in benzene (1:9), was used. The bleed-
Table 1.
ing column experiments utilized a pair of 1-meter columns packed with 20 weight % Tween 80 on 60- to 80-mesh Chromosorb W. Two sets of chromatograms were obtained on these columns. For one set a sample flow rate of 120 cc. per minute was used with the columns programmed from 100" to 305" C. at 7.9" C. per minute. Samples (0.032 ml.) yielded chromatograms with a pair of peaks at an attenuation of 2. A second set of chromatograms was obtained a t a sample flow rate of 60 cc. per minute with the columns programmed from 125"to 320°C. a t 7.9" C. per minute. A sample size of 0.008 ml. was used to give a pair of peaks at a n attenuation of 1. Under both sets of conditions, the naphthalene was eluted below 175" C. where the Tween 80 substrate was not bleeding appreciably and the p-terphenyl was eluted at 305" C. where the single-column baseline would just have gone off-scale at an attenuation of 1 on a 1-mv. recorder. The same binary mixture also was chromatographed on a less volatile column packing where both components were eluted below the temperature a t which this packing began t o bleed appreciably. The nonbleeding column experiments utilized a pair of 1-meter
Results of Quantitative Study
Ratio of Peak Areas (Naphthalene to p-Terphenyl) Sample Column Detector Bleeding Tween 80 Columna Nonbleeding Apiezon L Columns Flow Signal No. of No. of Rate, Attenua- ObserObsercc./min. tion vation Mean U vations Mean U 120 60
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columns packed with 25 weight yo Apiezon L on 30- to 60-mesh Chroinosorb u'. Two sets of chromatograms were obtained on these columns to correspond to those on the bleeding columns. The same flow rates, temperature programs, and sample sizes were used. Under these conditions the naphthalene was eluted a t 175" C. and the p-terphenyl below 320" C. where a single-column base-line would not have drifted up-scale more than 10% a t an attenuation of 1. Six to eleven chromatograms of the binary mixture of naphthalene and p terphenyl were obtained at the two flow rates on both the bleeding and nonbleeding columns. The naphthalene and p-terphenyl used in making the mixture were each checked for purity on both columns to make certain that no minor impurity in either component was being resolved on one column and
counted with a major coinponent on the other column. The results of the study are given in Table I in terms of the averages of the rat'os of the two component peak areas on each of the two columns. Comparison of the average peak area ratios a t each individual flow rate shows very excellent agreement indicating that there is no appreciable change in the relative response for a component eluted when the column is bleeding to the extent studied here. This conclusion applies to thermal conductivity detectors; quite different results might be obtained if detectors with more limited linear response ranges were used. The 8% difference in the peak area ratios a t the two different flow rates cannot be explained entirely by the effect on relative response of the quite different flow rates and sample sizes. If the flow controller on the sample side
reproducil)ly failed to maintain the flow a t 120 cc. per minute during the entire temperature program, a peak area larger than expected nould be obtained for the p-terplienyl and would lend to the ~ i p parent difference in peak area ratios which was observed. ACKNOWLEDGMENT
The authors acknowledge the assistance of L. M. Olszewski in obtaining some of the chromatograms required for the quantitative study. LITERATURE CITED
(1) Emery, E. M., Koerner, 11'. E., AXAL. CHEM.33, 523 (1961). RECEIVEDfor review March 8, 1962. Accepted July 13, 1962. Presented in part at the Symposium on Gas Chromatography, Divisions of Analytical and
Petroleum Chemistry, 139th Meeting, ACS, St. Louis, Mo., March 1961.
Gas Chromatographic Response as a Function of Sample Input Profile CHARLES N. REILLEY, GARY P. HILDEBRAND, and J. W. ASHLEY, Jr. Department of Chemistry, The University o f North Carolina, Chapel Hill, N. C.
b A series of equations based on a simple principle is developed to describe the shape of chromatographic curves in terms of the mode of sample injection. This principle is used to develop useful parameters to describe step chromatography curves, and comparison is made to the corresponding parameters of impulse chromatography curves. The use of various input functions leads to many novel response shapes, thus considerably broadening the scope of gas chromatographic techniques. Applications of various sample input profiles to quantitative analysis, preparative chromatography, the study of column phenomena, and the monitoring of industrial process streams are discussed.
terpret. Also, only one component of a mixture could be isolated in pure form, so the technique, understandably, did not find wide use as a preparative tool. Therefore, its major use has been confined to the determination of adsorption isotherms (6, 7, 11, 12). By coating an inert solid support with a partitioning liquid and by using sufficiently dilute solutions of solute in the carrier gas, as suggested by Boeke (1, d ) , linear isotherms can be obtained, a
and introduction of sample in the form of a step becomes a feasible technique, In any chromatographic experiment the type of chromatogram obtained (the response) depends on the shape of sample input profile (the input function). Although the number of possible input functions is almost unlimited, a few interesting types are shown in Figure 1. In Curve A , the sample is introduced as a step function-i.e., the concentration of sample
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introduced by Tiselius (13, 14) and developed by Claesson (6),has not been exploited in gas chromatography as fully as its counterpart, peak elution analysis. Undoubtedly, the reason that little enthusiasm has been shown for the frontal technique lies in the fact that early experimenters employed solid adsorbants, which exhibit interdependent nonlinear isotherms. As a result, frontal chromatograms of multicomponent mixtures were difficult to inRONTAL
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