Preparation of Porous Layer Open Tubular Columns by Dynamic Coating and Rapid Conditioning Max Blumer Woods Hole Oceanographic Institution, Woods Hole, Mass. 02543
Porous layer open tubular (PLOT) columns offer many advantages over packed or wall-coated gas chromatogram columns ( I ) . However, difficulties in their preparation and the expense of commercial columns have limited their application. Recently, Nikelly (2, 3 ) has demonstrated that a simple dynamic coating procedure produces PLOT columns of excellent efficiency. His method centers around the use of suspensions of finely ground (less that 2 X 10-4cm) and sized diatomaceous earth, such as JohnsManville Chromosorb W, in a solvent containing the liquid phase. The preparation of this substrate is simple but time consuming; if larger particles remain, clogging of the column may occur during coating. A commercially available hydrophobic silica of colloidal dispersion, Silanox 101 (Cabot Corporation, Billerica, Mass. 01821) permits the easy preparation of PLOT columns that are efficient, rugged, condition more rapidly than any existing GC columns, and that accept a wide range of liquid phases. Silanox 101 is described as hydrophobic, fumed silicon dioxide with a primary particle size of 7 x lO-?cm. Apparently, it is formed by a vapor phase reaction, and it is subsequently trimethylsilylated. Silanox 101 forms stable suspensions in many solvents, and in dispersion with organic binders it adheres to metal, glass, and plastic surfaces and forms thin but mechanically surprisingly stable films. These properties are responsible for the excellent performance of Silanox in PLOT columns.
EXPERIMENTAL A p p a r a t u s . Two types of gas chromatographs have been used, the Models 600 D and 1200 of Varian Aerograph. T h e instruments are equipped with flame ionization detectors and are connected to 1-mV recorders of YS- and 1-second response. Neither capillary injectors nor stream splitters are used. T h e Ys-in. 0.d. columns are directly fitted t o t h e injectors; suitable Swagelok adapters are used for 41e-in. 0.d. columns. A capillary pigtail is fitted by means of a drop of self-curing silicone rubber into the lumen of the exit end of those capillaries used in the Model 600 D. This pigtail reaches into the center of the detector base. T h e Model 1200 instruments use the commercially available low dead volume adapter a t the detector. Here, also t h e commercial make-up gas adapter is used. This is not available for the older Model 600; instead sufficient helium for detector flushing and for best flame performance (about 10 ml/min) is mixed with the hydrogen external to the chromatograph. Type 304 and 316 stainless steel tubing was used, both as new a n d as reclaimed columns. Columns were prepared in these dimensions: h 6 - and YE-in. o.d., 0.02-, 0.027-, 0.029-, and 0.030-in. i.d. and approximately 12, 22, 33, and 70 ft in length. Procedure. New or used columns are rinsed with a t least 6 colu m n volumes each of methylene chloride, acetone, pyridine, water, 30% nitric acid, water, pyridine, acetone, and methylene chloride. The columns are then dried with nitrogen, inserted into a GC oven, b u t not connected t o a detector, heated t o 280 "C a t a helium f'low of 20 ml/min, and treated with several 2-pl injections of silylating agent, such as Silyl-8 (Pierce Chemical Company, Rockford, Ill.). For a typical coating mixture 0.2-0.5 gram of liquid phase is dissolved in 10 ml of methylene chloride. Chloroform or a mixture ( I ) R. Kaiser, Chrornatographia, 1-2, 34 (1963). (2) J. G. Nikelly, Anai. Chern.. 44, 623 (1972). (3) J. G. Nikelly, Anal. Chsrn., 44, 625 (1972).
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of chloroform with methanol ( 8 : i ) may also be used. This solution is added to 0.4-0.65 gram Silanox 101 in a vial with Teflon lined screw cap. T h e suspension is dispersed by brief (30-sec) immersion into a n ultrasonic bath. T h e performance of the suspension and of the resulting column can be judged by immersion into the mixture of a clean and polished metal spatula, followed by rapid withdrawal and drying. A thin, smooth, and mechanically resistant coat should be formed; a granular coat indicates insufficient dispersion and would result in lowered column efficiency; a heavy coat t h a t dries with cracking and crazing suggests too high a substrate concentration or viscosity; adhesion of the coating would be poor and the column might be clogged during drying. Most columns were coated according to (2, 3 ) , with air or nitrogen a t pressures to 100 psig, depending on column dimensions and viscosity of the coating mixture, a t linear flow rates of 10-30 cm/second. First, the coiled tubing is filled with the solvent used in preparation of the coating mixture. If Nikelly's coating apparat u s (2) is used, excess solvent is drained from the filling tube; flushing the column with gas is not necessary. Five- to ten-column volumes of suspension are then added to the filling tube, and the solvent and excess suspension are flushed through the column a t the desired flow rate. No constrictor is used on the column exit; a small bleed upstream of the coating apparatus permits a reduction in gas pressure when the last coating liquid emerges from the column. Afterward the gas flow is maintained a t 20-50 ml/min, until the odor of the solvent is no longer noticeable. Some columns were prepared by injection, first of the solvent and then of the coating liquid from a 20-ml Luer-Lok syringe into a column sealed a t the upstream end with a Swagelok fitting containing a silicone rubber septum. After drying, t h e column is mounted in the chromatograph but not connected to the detector. T h e helium flow is set a t about 20 ml/min and the column temperature is programmed a t 4 "C/min from 70 "C to near operating limit of the liquid phase. Nonpolar columns, such a s Apiezon L which do not react with silylating agents are then treated a t 240-280 "C by two injections of 1 pl each of Silyl-8. Next, the helium flow is adjusted for optimum column efficiency (2-3 ml/min) a n d the column is connected t o the flame detector. Sample Introduction. Because of the large injector dead volume special procedures are used for sample introduction: a . Solutions used in temperature programmed G C are injected a t a temperature near the boiling point of the solvent. Thus, injection of 1-2 p1 of CS2 a t 40-50 "C is tolerated without a n adverse effect on the column. Probably, the great dilution of the solvent vapor in the large injector prevents condensation in the colu m n which might occur if a smaller injector were used. After sufficient time for flushing of the injector, the column temperature may be programmed, or raised stepwise to a higher constant temperature. b. For constant temperature operation, as required in t h e determination of plate efficiencies, the gas volume passing through the column in the course of a separation must be large relative t o the injector dead volume; correspondingly. a relatively low operating temperature is required. The best operating temperature is determined by measuring apparent plate efficiencies a t several decreasing temperatures; the true column efficiency is a p proached asymptotically. All column efficiencies reported here have been determined in this manner, with n-alkanes and n-fatty acid esters for unpolar or polar liquid phases, respectively. c. Solid samples or w r y dilute solutions can be handled by deposition, dry or from a solution, a t room temperature on the outside of a glass rod, which is inserted into the injector. Any solvent present is evaporated before the insertion of the rod, therefore the column temperature may be low, and sufficient time may be per-
mitted for the distillation of the sample into the column ( 4 ) . Linear temperature programming or stepwise elevation of the column temperature produces excellent chromatograms, even from large volumes of dilute solutions and with better accuracy than possible with most stream splitters.
RESULTS AND DISCUSSION The Silanox films and suspensions differ from those of other silicas primarily because of their small particle size, their great hydrophobicity, and because of the absence of open pores. The diatomaceous earths used by Kaiser ( 1 ) and Nikelly (2, 3 ) exceed Silanox by two to three orders of magnitude in particle size. Because of its large surface area, Silanox stabilizes liquid phases more readily than diatomaceous earth. The absence of internally and partially blocked pores retains the liquid phases in intimate contact with the gas stream and explains the low HETP, the rapid drying, and the rapid conditioning of the Silanox columns. Silanox 101 forms thixotropic gels a t low solids concentration, especially in unpolar solvents. This is no disadvantage since adequate wall coverage with liquid phase can be obtained with dilute, and quite fluid suspensions. The high concentration of diatomaceous earth (12-30%) required previously for adequate loading with liquid phase may lead to plugging during coating. This is not experienced with properly prepared Silanox suspensions. The viscosity of Silanox dispersions in unpolar solvents increases with the duration of mixing or with the shear used in that process. Concentrations above 7% congeal even during moderate mixing, but the addition of a polar solvent, e . g . , methanol, counteracts this to some extent. Adequate dispersion is necessary and is most easily reached by short ultrasonic treatment. In fact, column plugging was regularly encountered when this step was omitted. The Coating Mixture. Nikelly discusses the variables t h a t are to be considered in preparation of the coating solution: plug size, choice, and concentration of solvents (plug fluidity), and concentration of the liquid phase. With diatomaceous earth, the plug size should not exceed the column dead volume; larger plugs are reported to cause intermittent flow and clogging. This is not observed with Silanox suspensions, nor is it necessary to precoat the column with stationary phase; a simple wash with the solvent component of the coating mixture is adequate and the solvent may be displaced directly by the coating mixture. A large plug (5-10 column volumes) is preferred for displacing the solvent in order to arrive a t a coating that is not diluted by the wash solvent. The addition of wetting agents, which are commonly used in the preparation of nonpolar columns such as Apiezon L, is not required. This illustrates the superiority of the coating with Silanox; omission of the wetting agent is very desirable especially if accurate retention indices are to be obtained for a particular liquid phase. Table I demonstrates the dependence of plate efficiency and column capacity on the composition of the coating liquid. Low liquid phase loading, such as in columns 1, 2 , and 7, leads to efficient columns of moderate sample capacity. The higher H E T P in column 7 is due to its larger i.d. of 0.031 in. In spite of this wide bore, the column exhibits excellent efficiency. Comparable columns of different length (1 and 2 , 8 and 9) have a similar H E T P , and this is expected in view of the low flow resistance; it demonstrates also t h a t the separating efficiency is not adversely affected by the unconventionally large injector volume used in this work. (4) M. Ehrhardt and M . Blurner, Environ. Pollut., 3, 179 (1972).
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Increasing the amount of liquid phase (columns 3, 4, and 5 ) lowers the efficiency while raising the sample capacity. Obviously, a modest increase in column length can regain the loss in resolution. Column 5 may have been overloaded with liquid phase, relative t o the amount of Silanox, but the H E T P and the performance are tolerable. A further increase of stationary phase and substrate (column 6), possible only with the addition of methanol to the solvent, leads to a poor column with excessive HETP. Upon drying, the wall coat cracks and deteriorates. A wide range of liquid phases may be coated in the same manner as Apiezon L, and columns of comparable efficiency result (columns 10-13). The excellent polar substrate FFAP yields a higher HETP, here as in packed columns. However, excellent separations are obtained and the lower efficiency is easily compensated by an increase in column length. Silicone rubber columns (OV-101 and SE-54) were prepared; however, the coating solutions, unless very dilute, are viscous and the resulting columns tail (OV-101) and offer little advantage over those described. Coating Procedure. Coating columns with Silanox suspensions is remarkably uncritical. A sufficiently well dispersed suspension is stable and no stirring of the mixture in the reservoir or tapping of the column during coating is required. Air or nitrogen have been used to pressurize the filling tube, with no noticeable difference in the resulting column. Coating rates varied from 10 to 30 cm/sec. On one occasion (column 2), a defective valve permitted the pressure during coating to rise abruptly beyond 160 psig; in spite of the excessively high and uncontrolled coating rate, an excellent column with a H E T P of 1.04 mm resulted. Clogging at high flow rates, reported by Nikelly, was never observed. Two columns were coated by syringe injection through a silicone rubber septum, first of the solvent, then of the coating suspension. Finally, the excess suspension was blown out, using a dry syringe, and the columns were dried with compressed air. The resulting columns (8 and 9) are not much inferior t o those coated conventionally. It should be possible to formulate stable coating suspensions that may be injected into empty tubing from aerosol type cans. A water-proofing aerosol formulation of Silanox 101, made available to us by Cabot Corporation, contains 1% of Silanox 101 in isopropanol with 0.7% of a silicone binder. This was discharged through a clean 12foot capillary that was then air dried and conditioned. The resulting column resolved completely a test mixture of C14-Czo alkanes with slight tailing of the peaks. The presence of several branched alkanes, between the n-alkane peaks, was readily apparent. Column Conditioning. The ease of drying and conditioning of Silanox coated columns offers a spectacular improvement over conventional PLOT columns. Drying of a coated column requires only a few minutes, compared to 1 / ~hour for Chromosorb W coated PLOT columns. Conditioning can proceed a t 4-6 "C/min from 70 "C to the upper operating limit of the column. In contrast, conventional PLOT columns have to be programmed a t 0.1 "C/ min; this must be followed by a n isothermal stay a t elevated temperature for 2-4 days ( I ) . Other workers have recommended that this initial conditioning be followed by a carefully selected routine of small injections to complete the column conditioning. None of this appears necessary for Silanox PLOT columns. Apiezon L columns silylated after a single rapid program are fully operational within as little as 70-90 minutes after the beginning of the coating step. Similar 982 * ANALYTICAL CHEMISTRY, VOL. 45, NO. 6, M A Y 1973
rapid conditioning is possible for other chemically stable liquid phases of low volatility, such as Carbowax 20 M and FFAP. In the case of more volatile liquid phases (e.g.. Emulphor 0), the initial program has to be repeated, but a higher programming rate may then be used. Only stationary phases which undergo chemical changes during conditioning, such as silicones, may require a more complex conditioning program. Heat treatment and silylation of the empty tubing before coating appears to give more efficient performance and reduced tailing, especially of reclaimed tubing. Loss of substrate during drying, due to failure of adhesion of the coating, was observed in several columns not so treated, but never in heated and silylated tubing. Sample Capacity. In terms of sample capacity, the Silanox columns correspond closely to the Chromosorb WPLOT columns (2, 3 ) . N-alkane samples of increasing size were chromatogramed a t a program rate of 4 "C/min. The first indication of peak broadening was used as a measure of the sample capacity of the columns (Table I). However, the column capacity is a complex parameter and difficult pairs may often still be resolved a t a load that causes considerable peak broadening ( 5 ) . The expected dependence of the capacity on the liquid phase content of the coating mixture is found (columns 1 and 5 ) and suggests flexibility in the tailoring of coating solutions for the need of high resolution or high sample capacity. However, with very high substrate or liquid phase loading, the deposited film deteriorates upon drying and the sample capacity suffers (column 6). At this time, almost forty PLOT columns have been prepared from Silanox 101. Sufficient information on the durability of the columns is not. yet available. However, the low reactivity of Silanox and the adequate wall loading with liquid phase suggest that the life expectancy should be comparable to that of other capillary columns. The use of Yg-in. 0.d. capillary PLOT columns and of the injection techniques described here, permits the simple conversion of many older gas chromatographs to capillaries. Even moderately short columns (10-20 f t ) reach or surpass the efficiency of packed columns; they offer unique advantages in preparation and in the predictability of the separations. CONCLUSIONS Silanox 101, a colloidally dispersed hydrophobic silica is an ideal support for the preparation of porous layer open tubular columns. The column performance is predictable, coating is simple, and the rapidity of conditioning is unequalled by any other gas chromatographic columns. Note added i n proof. During review, I became aware of a similar effort, using Silanox 101 and GE-SE 30. Readers interested in silicone rubber PLOT columns may wish t o consult the paper by A. L. German, C. D. Pfaffenberger, J-P. Thenot, M. G. Horning and E. C. Horning, Anal. Chern., 45,930 (1973). ACKNOWLEDGMENT I thank the Cabot Corporation for samples and P. R. Tully and J. Roach for advice on the use of Silanox 101. Received for review November 13, 1972. Accepted January 5 , 1973. This work was supported by the National Science Foundation (Grant GA 35646) and by the Office of Naval Research (N0014-66, Contract CO-241). Contribution No. 3079 of the Woods Hole Oceanographic Institution. (5) L. S. Ettre, J (1966).
E. Purcell, and K. Billeb, J. Chromafogr., 24, 335
