VOLUME 33, NO. 10, OCTOBER, 1956
GAS-PHASE CHROMATOGRAPHY JOHN R. LOT2 and CHARLES 8. WILLINGHAM Mellon Institute, Pittsburgh, Pennsylvania INTRODUCTION
The technique of chron~atographyhas in recent years become a powerful analytical and research tool. The principles of chromatography are simple, its application usually does not require elaborate apparatus, and frequently separations can be effected which are difficult or impossible by other methods. ITntil recently, chromatographic methods were limited to the handling of liquid mixtures or solutions. Although the phenomenon of gas adsorption on solids was well known and had found application to some analytical problems, this technique was not widely developed. In 1941 Martin and Synge (I) introduced liquidliquid partition chromatography, and suggested that separation of volatile substances should be possible if the moving liquid phase were replaced by a moving inert carrier gas containing the sample. Their suggestion was neglected until James and Martin (3) showed that mixtures of aliphatic acids could be separated by this method. Since that time, progress in this field has been extremely rapid. Numerous papers have been published, symposia have heen held, and several manufacturers have instruments in production.
Gas-phase chromatography' differs from the more familiar liquid-phase method in that the mixture to be separated is vaporized into a stream of inert carrier gas and passed through a column appropriately packed for separation by gas adsorption or gas-liquid partition chromatography.' A detecting device a t the exit end of the column indicates the presence of the separated components of the mixture as they emerge. In gas adsorption chromatography (3, 4, 5), the column is packed with a solid such as activated carbon, alumina, or silica gel. The sample is adsorbed and its components are selectively removed by elution by the carrier gas (elution chromatography) or by displacement in which the carrier gas contains a vapor which is more strongly adsorbed than the sample (displacemcnt chromatography). NOTE ADDEDIN PROOF: I n conformity with the agrccmcnt reached in the discussions of this topic a t the Symposium on Vapor-Phase Chromatography before the Division of Analytical Chemistry a t the 129th Meetingof the American ChemiodSociety. Dallas, April, 1956, the term "gas phase ehrom&ography" should he replaced by the preferable "gas chromatography." Similar preference should be given to "gas partition" in place of thr term "gas-liquid partition" where i t appoars in this text.
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Several papers dealing with the theoretical aspects of gas-phase chromatography have been published (2, 7). In gas-liquid partition chromatography, the potential method originally proposed by Martin and Synge, the sample components are alternately dissolved in and vaporized from a high-boiling liquid which is impregCorrler GO.
spectroscopy, but should prove to be rt valuable supplement to these methods. Although its main use thus far has been in analysis, gas-phase chromatography should find many applications in other phases of research work. The determination of partition coefficients(15) may be cited as just one example. This paper is intended not as an exhaustive review of gas-phase chromatography, but rather as an introduction to the subject. Sufficient literature references have been given, however, to serve as a point of departure for further study. Much of the research in this field to date has been carried on in industrial laboratories. Because of its simplicity and relatively low costs, gas-phase chromatography offers attractive research possibilities for the college laboratory and a considerable increase in interest from that quarter may be expected. APPARATUS
nated on an inert solid support. Because of differences in their gas-liquid partition coefficients, the components of the sample are moved along the column a t different velocities, and hence emerge from the column at different times. The process is thus somewhat analogous to a small-scale extractive distillation. For low-boiling materials gas-adsorption chromatography gives good results, but gas-liquid partition chromatography is the preferred method for samples which are liquid a t or above room temperature and for strongly adsorbed substances which are difficult to remove from active adsorbents even a t elevated temperatures. When a sample has completely passed through a partition column, the column is in its original condition and can be used immediately for another sample. Because of irreversible adsorption, an adsorption column cannot be used indefinitely without regeneration. The use of both methods in the same column, by supporting a very small quantity of a partitioning liquid on active carbon, has been reported (6). The majority of work done in gas-phase chromatography has used the gas-liquid partition technique, and for that reason most of the present paper will deal with this type of separation. Gas-phase chromatography as an analytical tool could be applied to nearly any material which has an appreciable vapor pressure. Separations of mixtures of such diverse types as acids (8, 7), alcohols (a), amines (9, lo), esters (8, l l ) , and several classes of hydrocarbons (6,12, IS, 14) have been reported. It is rapid, uses very small samples (with the possibility of recovery of the sample components for examination by other methods), and requires fairly simple and comparatively inexpensive apparatus: ~ i k eany other technique, gae-ihase chromatography has li&itations. I t is not likely to render obsolete such well-established methods as analytical distillation and infrared or mass
A block diagram of the apparatus required for gasphase chromatography is shown in Figure 1. The separate components will be discussed below. Columns. Since the column handles only gases, it may be operated in any position. Long columns may be bent or coiled into any convenient shape. Although glass columns are frequently used, metal tubing has the advantages of ruggedness and ease of connection to other parts of the system. Soft copper tubing is perfectly satisfactory provided it is not attacked by the sample vapors; stainless steel is more resistant, but is somewhat more difficult to shape. The type of packing for the column will depend upon whether adsorption or partition chromatography is under study. As bas been mentioned in the introdnction, active adsorbents such as carbon or silica gel are used in the former method, while inert solids impregnated with high-boiling liquids comprise the packing for the latter operation. Diatomaceous earth (Celite) has been a popular supporting material, but other substances, such as crushed and screened firebrick (16) and glass spheres (17), have been used. Esters, such as phthalates; mineral oils; vacuum-pump oils; silicones; and higher alcohols are some examples of the many liquids which have been tested in partition columns. If the sample components form ideal solutions with the liquid phase in the column, they will be eluted by the carrier gas in order of decreasing vapor pressure (or of increasing boiling point). If nonideal solutions are formed, the sample components may be held back to an extent depending upon the deviation from ideality. This may of course prove to be an advantage in some separations. The column liquid should have a negligible vapor pressure a t the temperature of operation, it should be stable, and it should not react with the sample. Further discussion of the properties of column liquids has been niven bv Hausdorfi (18). ~, At present the choice of a column liquid is largely ~
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VOLUME 33, NO. 10, OCTOBER, 1956
empirical, with each laboratory exploiting those liquids which it has found to give acceptable separations. It is obvious that no one class of liquids will be satisfactory for the separation of all types of mixtures, but it is likely that columns that combine the effectiveness of several liquids might be of wide applicability. This field is a very active one a t present, and many new developments may be anticipated. Detectors. In liquid-phase chromatography the column effluent may be divided on the basis of color or other properties, or fractions may be collected on a time or volume basis and analyzed by appropriate methods. In gas-phase work, a continuously operating sensing device is used, and its signal is applied to a strip-chart recorder. As the sample components emerge from the column, their concentration in the carrier gas stream is quite low (of the order of a few per cent) and consequently a sensitive detector is required. Any property sensitive to small changes in concentration might be used as the basis for a detector, but only a few have proved practical. In some of the earliest work in this field, James and Martin (2) studied fatty acids and amines by means of an automatic recording titrator. Such a detector would of course be suitable only for ionizable samples. James (19) described a gasdensity balance which gave good results, but this does not appear to have found widespread use by others. Among other detection methods might be mentioned the measurement of change in surface potential between two metal plates exposed to the column effluent (20) and changes in temperature of the flame of burning carrier gas (hydrogen) caused by the presence of sample (21). Although not a detector in the usual sense, the method devised by Janak (88) is interesting: carbon dioxide is used as the carrier gas, and after passage through the column the carbon dioxide is absorbed in concentrated potassium hydroxide solution and the volume of the insoluble sample components is measured. In this country, most detectors are based on the difference in thermal conductivity between the pure carrier gas and the carrier gas containing sample vapor. In essence, a diierential thermal conductivity cell is a Wheatstone bridge in which carrier gas flows over or near two of the resistors (thermistors are sometimes used) and the column effluent over or near the other two. In some cells only one resistor is exposed to carrier gas and one to column effluent, and external fixed resistors make up the other two bridge arms. The cell resistors are heated by passage of current, and the bridge is adjusted so that it is in balance when only carrier gas is coming through the column. Emergence of sample from the column causes a change in the thermal conductivity of the effluent as compared with the carrier gas. This results in a change in temperature, and consequently a change in resistance, of the resistors exposed to the column effluent. The resulting- unbalance of the bridge is then applied to the recorder. Thermal conductivity cells are available from several
Three-foot column, 25-C., NI aarrier gsl. 5.7 ml./min. now rate thermistor-type thermal aonduotivity detector.
manufacturer^,^ but their construction is not beyond the ability of a good technician (17). Typical wiring diagrams and a discussion of cell sensitivity have been published (16). Thv rrquirrments for a recorder suitalk for use with a thrrnml conducti\.itv cell are not nurrivulrlrlv strinrent. Probably any such instrument having a full-scale sensitivity of ten millivolts or less could be used. In theory, the output of the cell could be fed to a galvanometer and r e a d i i taken manually, but this would he far less convenient and less accurate than the use of a recorder. Carrier Gas. If a differential thermal condnctivity cell is used as a detector, the carrier gas should have a thermal conductivity as different as possible from that of the samples. Hydrogen, whose thermal conductivity is of the order of seven times that of most organic vapors, is best in this respect. Hydrogen does, in fact, give excellent results, hut its use is attended by some hazard. and the nossibilitv of reaction of unsaturated > compounds with hydrogen on the hot cell filaments should be considered. The latter objection is of course Z important only if the Sam- 5 :: plea are to be recovered. Helium gives a sensitivity 8 second only to hydrogen. In addition, helium is safe to handle and cannot react with samples. Its cost is eo 10 o T i m e , Minutes somewhat greater, but not prohibitively so. Where ~igure 3. %parstion of 1.0highest sensitivity is not merk Hexenms-F&ev Flow Rate reauired. nitronen is satisConditions same as thoae for and even 'lean air Figure 2 except Row rate of 30 could be used. However ,~./,i,.
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Six-foot column, 60°C., Nz aarrier gas, 11.0 rnl./min. flow rate, .*hot wire"-type thermal conduotivity detector.
nitrogen, unlike hydrogen or helium, does not give the same response for all types of samples. This is not serious for qualitative work. Calibration with known amounts of pure samples is necessary for quantitative analysis. Sample introduction. Gaseous samples are easily introduced into a gas-phase chromatographic system by sweeping carrier gas through the sample bulb. A pair of three-way stopcocks permits by-passing the bulb until the sample is to be transferred to the column. Liquid samples have most often been introduced by some type of hypodermic b " syringe through a rubber dz serum bottle cap. James and Martin (2) have used ultramicropipets with good results. Dimbat, Porter, and Stross (16) have designed a system to be used 0 with weighed samples in 15 10 5 ~ iM ~ ~~U I ~ ~S . sealed glass bulbs. AlF i w - 8. Sspa-tion of though considerably more m u i c H...nss-Hi.her Ternpmr.tura elaborate than those alr e a d y mentioned, t h i s Conditions same as thoas for Figure 4 except temperature of give reIOO'C. sults.
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Figures 2, 3, 4, and 5 show some typical results obtained in the authors' laboratory. The sample in all cases was an equal-volume mixture of the five isomeric hexanes, and the columns were packed with a mixture of 40 per cent di-n-decyl phthalate and 60 per cent Celite 545. The "balanced-column" flow system (14) was used. I t should be noted that only four peaks are shown in these figures. Two of the hexanes, 2-methylpentane, and 2,3-dimethylbutane have nearly equal times off the column under the conditions used, and are therefore not separated. These compounds coming off together yield the large second peak shown in the figures. Separation of this pair, using slightly different conditions, has been reported (6, 16). It is evident from inspection of Figures 2 and 3 that the effect of flow rate, a t least within these limits, is merely to compress or expand the time axis. A similar effect could be had by changing the chart speed of the recorder. At first sight it might appear that the lower flow rate gives an improved separation. Closer inspection of the curves will show, however, that in Figure 2 the peaks are much broader and their centers are less distinct. Consequently, if time measurements are to be taken from the chart, it is more difficult to locate accurately the center of the peak. I n other words, the resolving power of a given column operated at a given temperature is not a function of the flow rate. Comparison of Figures 4 and 5 shows that increased temperatures, like increased flow rate, shorten the time requked to move the sample through the column. Higher temperatures, however, definitely decrease the resolving power of the column, as is shown by the disappearadce of one peak in the 100' run. Thus for maximum separation the column should be operated a t the lowest possible temperature. If the low temperature results in an inordinately long time for the sample to travel through the column, this condition can be corrected to some extent by increasing the flow rate. For samples with wide boiling ranges, it may not be possible to examine the entire sample using a single set of operating conditions and several runs may be necessary. I n general, the best conditions for any given apparatus and sample must be determined by trial and error. When a particular apparatus is operated a t fixed conditions, the time required for a given substance to reach the end of the column is characteristic of that substance. This time, the retention time, is a function of temperature, flow rate, carrier gas, and column packing, as well as dimensions of the apparatus. Therefore, individual calibrations are required. It is possible to eliminate the effect of flow rate by expressing results in terms of retention volumes (product of retention time and flow rate). It is convenient t o express retention times or volumes relative
VOLUME 33, NO. 10, OCTOBER, 1956
to some standard material, thus eliminating all variables SUMMARY except temperature. For example, retention volumes Gas-phase chromatography offers a simple and rapid and relative retention volumes compared to n-pentane method for the separation and analysis of nearly any liave recently been published for the hexenes and mixture whose components have appreciable vapor hexanes (14). pressures. Its apparatus requirements are relatively Therefore, if the apparatus has been calibrated with simple and inexpensive, and the technique of operation known compounds, identification of an unknown may is easily mastered. often be established by comparison of the unknown's LITERATURE CITED retention time or retention volume with those of the ( 1 ) MARTIN,A. J. P., AND R. L. M. SYNGE, Bioehem. J. (Lonstandards. don), 35, 1358 (1941). Various relationships involving retentionvolumes have A. T.. AND A. J. P. MARTIN.Biochem. J. (London). (.2.) JAMES. SO, 679 (1951). been discovered, and these may be of value inidentificaD. H., AND C. S. G. PHILLIPS,J. Chem. Soc., 1953, ( 3 ) JAMES, tion work. For example (7), a plot of the logarithm of i..finn . .. retention volume against number of carbon atoms gives (4) PATTON, H. W., J. S. LEWIS, AND W. I. KATE,Anal. Chem., a different straight line for each homologous series. 27, 170 (1955). ( 5 ) RAY,N . H., J. Appl. Chem. (London), 4 , 8 2 (1954). Again, a plot of retention volumes in one column packF. T., H. S. KNIGHT,AND S. GROENNINM, ( 6 ) EGGERTSEN, ing against those in another gives a series of straight J. Appl. Chem. (London), 28, 303 (1956). lines for diierent classes of compounds (7). A. T.. AND A. J. P. MARTIN.AnalwL 77. 915 ( 7 ) JAMES. If operation is such that the response of the detector is independent of the nature of the sample, the relative areas of the peaks on the recorder chart are proportional to the mole percentages of the components in the sample. This gives a ready method for quantitative analysis. 4. Hemoon, Nature, 172,1101 (1953). B. W., D. E. CRALKLEY, AND D. HARVEY,J. Peak areas may be measured with a planimeter, or (12) BRADFORD, Imt. Petroleum, 41, 80 (1955). estimated from the product of their height and width D. H.. S. A. FLECK.AND F. H. BUROW. Anal. 113) LICHTENFELS. a t half-height (B). The sample size does not enter . . C h m .,. 27.'1510 ll955). . ,~ into the calculation and therefore the sample need be ( 1 4 ) SULLIVAN, L. J., J. R. LOT., AND C. B. WILLING^, Anal. Chem., 28, 495 (1956). only of such size as to yield peaks which can be easily measured. If, however, only one component is to be (15) PORTER,P. E., C. H. DEAL,AND F. H. STROSS,Anal. Chem., t o be published. determined in the sample, the sample size must be (16) DIMBAT, M., P. E. PORTER,AND F. H. STROSS, Anal. Chem., accurately measured and the apparatus must be cali28,290 (1956). brated with known amounts of the desired component. (17) CALLEAR. A. B.. AND R. J. CVETANOYIC. Can. J. Chem.. 33. 1256 (i955). ' Alternatively, a known amount of some internal standHAUSDORPP. H. H.. Anal. C h m . . to he ~ublished, ard may be added to the sample, and the percentage of JAMES, A. T., ~fg.'chemiat,26, 5 (1955): an unknown component calculated from a proportion GRIFFITBS, J. H., D. H. JLMES, AND C. S. G. P ~ L L I P S J., involving its peak area and that of the standard. Chem. Soc., 1954, 3446. SCOTT,R. P. W., paper presented a t Gas Chromatography Detector response nearly independent of sample Symposium, Ardeer, Scotland, May 20, 1955. nature is achieved with hydrogen or helium as carrier JANAK. J.. Chem. Listv. .. 47.. 464 (1953): C. A , . 48.. 3196 gases, but not with nitrogen. In the latter case, (195i). correction factors can be determined by measuring the CREMER,E., AND R. MULLER,Mikroehems uer. Mikroehim. peak area per unit quantity for different compounds. Ada, 36/37, 553 (1951).
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SIGMA DELTA EPSILON ANNOUNCES POSTDOCTORAL FELLOWSHIP SIGMADelta Epsilon, graduate women's scientific fraternity, is offering for the year 1957-58 a 51600 postdoctoral research fellowship to women specializing in the mathematical, physical, and biological sciences. Women with the equivalent of a Ph.D. degree, omying on research in the mathematical! physical, or biological sciences, who need finitncial assistance and give evidence of high abil~tyand promise, are eligible. During the term of her appointment the appointee must devote the major pwt of her time to the approved research project and not engage in other work for remuneration (unless such work shall have received the written approval of the Fellowship Awards Board). Application blanks may be obtained from Dr. Dorothy Quiggle, Petroleum Refining Laboratory, The Pennsylvania State University, University Park, Pennsylvania. These must be returned before February 1, 1957. Annonncement of the award will he made early in March, 1957.
