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I n our application, we allowed 25-OH-D3t o be deposited at t h e front because there were no other fast-moving radioactive components in our sample. Our procedure permitted optimal use of t h e available length of the T L C plate for separation of the more polar metabolites. Other applications may require additional chromatography t o distinguish fast-moving sample components. I n TLC, t h e rate of development and R, values may vary from one r u n t o t h e next. If development is controlled by co-chromatographing a visible marker so that it reaches a predetermined point on t h e chromatogram, continuous-development TLC is reproducible. Ideally, t h e marker should have a n R, value similar t o t h a t of t h e compounds t o be isolated. Methyl red was suitable for our application; however, a m u c h slower-moving dye (e.g., neutral red) would be preferable if longer development were necessary. T h e "CRC Handbook of Chromatography" gives t h e R, values of many dyes in several solvent systems (16). I n conventional TLC, t h e primary approach to increasing resolution is limited t o selection of a n appropriate solvent system. T h e technique described in this paper provides a n additional means with which t o improve resolution-the amount of solvent flow.
LITERATURE CITED D. L. Saundeys, Anal. Chem., 46, 470-473 (1974). L. R. Snyder, Principles of Adsorption Chromatography. The Separation of Nonionic Organic Compounds", Dekker, New York, 1968, p 208. L. R. Snyder and J. J. Kirkland, "Introduction to Modern Liquid chromatography", Wiley-Interscience, New York, N.Y., 1974, p 68. J. A . Thoma, Anal. Chem., 35, 214-224 (1963). J. A. Thoma. J . Chromatogr., 12, 441-452 (1963). J. M. Bobbit et al., "Introduction to Chromatography", Reinhold-Von Nostrand, New York, N.Y., 1968. M. Brenner and A. Neiderwieser, Experientia, 17, 237-238 (1961). N. Zollner and G. Wolfram, Klin. Wochenschr., 40, 1098-1101 (1962). E. V. Truter, J . Chromatogr., 14, 57-61 (1964). L. M. Libbey and E. A. Day, J . Chromatogr., 14, 273-275 (1964). R. W. Gray, J. L. Omdahl, J. G. Ghazarian, et al., J . Biol. Chem., 247, 7528-7532 (1972). J. C . Knutson and H. F. DeLuca, Biochemistry, 13, 1543-1548 (1974). G. Jones and H. F. DeLuca, J . Lipid Res., 16, 448-453 (1975). A. M. Rosenthal, G. Jones, S. W. Kooh, and D. Fraser, "Endocrinology of Calcium Metabolism", D. H. Copp and R. V. Talmage, Ed., Excerpta Medica, Amsterdam-Oxford, 1978, p 371. D. Kritchevesky and S. Malhotra, J . Chromatogr., 52, 498-499 (1970). G. Zweig and J. Sherma, "CRC Handbook of Chromatography", Vol 1, CRC Press, Cleveland, Ohio 1972.
RECEIVED for review December 8, 1977. Accepted J u n e 15, 1978. This work was supported by a grant (MA 5139) from t h e Medical Research Council of Canada.
Gas Chromatographic Injector Attachment for the Direct Insertion and Removal of a Porous Polymer Sorption Trap Harney Peterson Adolph Coors Company, Golden, Colorado 8040 1
Gary A. Eiceman, Larry R. Field,"' and Robert E. Sievers Department of Chemistry, University of Colorado, Boulder, Colorado 80309
In recent years, there has been a growing interest in t h e analysis of trace organic compounds in the environment (1-3). T h r e e of t h e most widely used methods t o t r a p and concentrate organic compounds from a n aqueous matrix have been: direct sorption of organics from water onto charcoal, followed by solvent extraction and concentration ( 4 , 5 ) ;direct liquid-liquid extraction, followed by solvent evaporation (6, 7); and, inert gas stripping of volatile organics onto polymeric resins, followed by thermal desorption (8-10). It is our belief t h a t the third method, despite its limitations, affords the best approach t o analysis of trace volatile organics in the environment. With this method, long pre-concentration times, extensive sample handling, a n d the use of large quantities of ultrapure solvents can be avoided. One major objection t o this method has been the complex and expensive hardware required t o thermally desorb a sample from a porous polymer t r a p onto a gas chromatographic column. Some researchers using this technique use expensive heated switching valves, heated desorption ovens, and heated sample transfer lines ( 1 1 , 12). Two previous attempts t o circumvent this hardware problem have been: first, removal of t h e septum a n d septum cap, direct insertion of the t r a p into t h e GC injection port, a n d replacement of t h e septum and cap; and, second, insertion of either sealed glass tubes or metal caps containing t h e polymer followed by destruction of t h e seal inside t h e injection port (13, 14). T h e s e techniques have disadvantages. Problems encountered in the first approach include backflushing of volatile Present address, Department of Chemistry, University of Washington, Seattle, Wash. 98195. 0003-2700/78/0350-2152$01 OO/O
compounds before t h e septum is resealed and limited desorption efficiency of trapped compounds due to poor carrier gas flow through t h e t r a p and injection port. .4severe disadvantage to the second method is that it is necessary to clean t h e polymer from t h e injector port following each analysis. I t was our intention t o devise a method in which most of t h e external hardware could be eliminated. In addition, we wished t o design and construct a device which would overcome t h e disadvantages listed for t h e previously used techniques.
EXPERIMENTAL Design and Construction. The two independent devices we have designed each offer different advantages to potential users. Injector attachment A (see Figure 1) requires little machining and virtually no modifications to the gas chromatograph (GC), while fabrication of injector attachment B (see Figure 2) demands considerable machining and some GC injector port alteration. Design A was developed for a Hewlett-Packard model HP5730A gas chromatograph in which the injection port internal diameter is -5 mm. The advantages of this design are its simplicity and minimal change of the instrument. Since each GC manufacturer has a different injection port design, the description of the injector attachment is general, and exact dimensions are not specified. The injector body (design A) is made from a 15.2-cm piece of 6 mm o.d. x 4 mm i.d. Pyrex tubing with composite Swagelok fittings attached a t each end (see Figure 1). On the end that attaches to the injection port, one half of a Swagelok inch-'/,-inch union is silver soldered to a l / inch Swagelok nut, the threads of which match those on the HGi730 injection port. Teflon ferrules are used throughout this design to provide air-tight seals. On the other end, a reducing Swagelok union I / , inch-1/8-inch allows a 0.3-cm o.d. brass rod to slide through a 1978 American Chemical Society
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Figure 1. Diagram of sorber cartridge injector design A. In addition to the injector system, the brass plunger rod and sorber cartridge are depicted separately
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Teflon ferrule. This rod is 17.8 cm long and has a loop handle bent in one end to facilitate easy insertion and removal of the sorption cartridge (see Figure 1). Onto the other end of this plunger rod, two prongs of 0 5 m m o.d. spring metal wire are soldered. Soft solder has been used so that spring temper in the wire is retained. One side of this spring is straight, while the other side has a small right-angle bend a t the end. This combination straight wire and hook arrangement snaps into a small hole in the sidewall of the sorption cartridge. This flexibly firmly affixes the plunger rod to the cartridge, thus permitting its easy insertion and removal. T h e rod slides through the Teflon ferrule to seat the sorber in the injection zone without interruption of carrier flow. There is no injection surge or peak. The sorption cartridge is considerably smaller than the one used in design B. Tubes have been made of either 3-mm o.d. Pyrex or of thin-wall 0.32-cm o.d. brass telescoping tubing. In either case, the bottom or inner end inside dimension of the sorber is reduced by fire polishing glass or by tapering the metal with a lathe, in order to hold a small plug of glass wool while permitting gas passage through the sorber. A small hole is drilled in the sidewall 4 mm from the other end of the sorber. This hole receives the spring hook arrangement described above. Tube length is determined by the length of the heated injection port, which varies among different manufacturers. The described sorbers are about 55 mm in length. T h e polymeric sorbent packing is drawn into the tube by vacuum and retained against the glass wool plug. A small amount of polymer is removed t o allow packing glass wool just below the side hole. The only instrument modification required for design A is the addition of another hybrid Swagelok fitting to match the oven union fitting and the diameter of the columns in use. This fitting supports the Teflon seat or seal. Above this, a Pyrex tube is cut so that it is long enough to occupy the full length of the injection port. This tube serves to guide the sorber to seat in the Teflon seal. T h e seal is made from a Teflon washer with an o.d. equal to the i.d. of the injection port. The i.d. of this washer is drilled by steps from just over 0.32 cm to enough under 0.32 ern t o act as a tapered seat into which the glass cartridge seats in the injected position. When the cartridge is seated in the Teflon seat, the carrier gas is forced through the cartridge and into the inlet end of the analytical column. T h e sorbent material most commonly used in our work is 80-100 mesh Tenax GC (-32 mg). However, 40-mesh XAD-2 and Carbopak C with a 0.170loading of S P 1000, and Porapak Q, have also been used as sample sorbing materials in these cartridges. T h e various sorbing properties and breakthrough volumes of these packing materials have been discussed previously in the literature (15,161. In addition to polymer packed cartridges, empty tubes have also been used as a solid injector for a narrow strip of polymeric beverage can liner. T h e various volatile compounds released during the flash heating of lining material give sharp chromatographic peaks with no appreciable band broadening. The small size of the sorbing cartridges used in design A enables the direct injection of these trapping tubes into the injection port without thermal focusing (thermal focusing is a technique whereby the organic vapors are condensed on the head of a cooled ( - 20
3C-32fiFT 31 -USE
Figure 2. Detailed drawing of injector design B. The sealing mechanism for sorber cartridge on t h e head of the analytical column is also demonstrated
"C) analytical column as they are thermally desorbed from the sorbing cartridges). T h e peaks obtained from' these sorption cartridges, corresponding to trace volatile organic compounds, give well defined Gaussian shaped peaks. This is not the case for the larger traps used in design B. Design B is illustrated in Figure 2. The body of the attachment is fabricated from 5.0-cm o.d. 4020 aluminum rod. The heat sink, machined from the same stock, is attached to the body by a 20/40 threaded couple. A Vitron O-ring, located at the thread connection in a compression groove permits adjustment of the O-ring diameter. A Varian model 2400 GC injection port was bored to 0.6-cm i.d. to accept the 0.6-cm 0.d. sorber cartridges used in this design. A 11/7 thread in the heat sink was used to attach the heat sink to the injection port in place of the septum cap. A septum, with its center bored out with a =1 cork borer, is placed between the injection port and the heat sink. Stainless steel 316 cartridges (7.6 cm long X 0.6 cm o.d. X 0.48 cm id.) were packed with 60-80 mesh Tenax GC porous polymer. Attachment to the plunger rod is made with a 1.9-cm piece of 6-32 screw stock silver soldered into the end of the cartridge. Thus, by screwing the cartridge attached screw into the corresponding female threads on the end of the plunger rod, it is possible to insert and remove the cartridge intact. This feature allows one to reuse the sorbing material and cartridges without subsequent cleaning of the injection port. Ineffective sealing between the trap and injector port is eliminated through the use of a simple pressure tight Vespel cup that rapidly forms a secure seal. After insertion, all of the carrier gas is diverted through a hole (3.0-mm diameter) located near the screw end of the cartridge. This should result in an enhanced thermal stripping efficiency during the flash desorption process, as compared to the loose tube insertion technique discussed above. T h e problem of backflushing the volatile compounds from the cartridge during the insertion step has been eliminated by keeping the sorption cartridge in the cool O-ring sealed portion of the injector body prior to its insertion into the heated injection port. During and after the cartridge insertion step, these same O-rings prevent backflushing or venting of the analyte components. The Vespel cup seat used to seal the cartridge into the carrier gas stream upon injection is made from a composite of both a '/,-inch and '/,-inch Vespel ferrule. Figure 2 shows the concentric arrangement (press fit smaller into larger) of these two ferrules as well as the flat machined surface of the inside '/d-inch ferrule. This arrangement forms the Vespel cup-like seat for the sorber cartridge. Procedure. T h e method for trapping trace volatile organic compounds from both air and water samples of various sorbing materials has been treated in detail by others (17, 18). T h e head-space sparging procedure of Hertz et al. ( 1 9 ) was used for concentrating volatile organics from water throughout this work. A similar approach was used for concentrating volatile organics from beer except that the natural effervescence of carbon dioxide gas was used and carbon dioxide functioned as the stripping gas in place of nitrogen. Air sampling was done by drawing known volumes of air (6-12 L a t 50 mL/min) through the sorbing cartridges. These cartridges were then inserted into the injector in such a manner that the flow of carrier gas through them was
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3. CI HC:C Cln
0
60
I20
Time ( m i d 5
--
Figure 3. Gas chromatographic profile of volatile compounds present in 400 m L of beer vapor. Injector: design A. Sorption material: Tenax-GC (60-80 mesh). Column: 50 c m X 0.64 c m 0.d. X 2 mm i.d. Pyrex, 0 . 1 YO w / w of SP-1000 on Carbopak C. Temperature program: 30-200 "C at 10°/min. Temperatures: 180 O C injector, 200 O C detector. Attenuation: 5 1 2 X . Carrier: N, 1 8 mL/min
reversed from the flow of sample gas during the sample collection step. Breakthrough studies were performed by placing two sorbing cartridges in series. In air sampling work, no prefilter was used ahead of the cartridge. Consequently, the chromatograms represent compounds from particulates and the vapor phase. In both injector attachment designs, the main body of the injector is screwed onto the injection port in place of the septum cap. Only the injector rod is removed for each analysis. Once the rod is removed, the sorbing cartridge is attached to the rod by either a hooking or screwing motion, depending on the design. Next, the plunger rod and attached cartridge are inserted into the injector body until the cartridge forms a seal. This seal is accomplished by the Teflon ferrule in design A and by the second O-ring in design B. The compression fitting design around this seal permits tension adjustment on the cartridge at this point. This adjustment ensures a good air-tight seal. A 60-s time period is allowed to elapse after the cartridge has been inserted into the main body. This enables the carrier gas flow to stabilize before the sample is injected. Next, the plunger rod is thrust into the heated injection port until the cartridge is sealed to the cup-like seat. The sealing is easy to detect as there is a slight resistance to the plunger rod thrust as the cartridge enters the cup seat. The smaller cartridge used in design A is quickly retracted after a 2-min thermal desorption period to the unheated main body of the injector. The larger cartridge used in design B is left in the inject position for 10 min, with the column oven cooled to -20 "C with dry ice, before it is withdrawn. By this thermal focusing technique, the volatile organic compounds which are thermally released from the cartridge are condensed in the inlet of the analytical column. Then the GC analysis is initiated by normal temperature programming.
RESULTS AND DISCUSSION T h e convenience of concentrating trace levels of volatile organic compounds in small sorbent-filled cartridges, combined with t h e ease of cartridge insertion, desorption, and removal, makes this injection device very useful. Two representative chromatograms are shown (Figures 3 and 4) to illustrate t h e excellent peak symmetry and absence of significant broadening characteristic of this type of cartridge injection. T h e precision with which retention time measurements can be reproduced using this injector system is demonstrated by triplicate analyses of C2, C4, and C6 methyl esters on four different sorbing cartridges. T h e percent relative standard deviations were found t o be zk3.9, *L6, and f0.9%, respectively. Figure 3 shows t h e chromatographic analysis of volatile components found in a fresh batch of commercially prepared
Figure 4. Typical gas chromatogram of volatile organic compounds found in Metro Denver waste water. Injector: design B. Sorption material: Tenax-GC (60-80 mesh). Column: 30 mm X 1.5 mm 0.d. X 0.25 mm i.d. capillary coated with OV-101. Temperature program: -20 "C-250 OC at 2 "C/min. Temperatures: 250 OC injector, 250 "C detector. Attenuation: 6 4 X lo-'*. Carrier: N, 0.5 mL/min
beer. This chromatogram was obtained with injection device
A, without the aid of thermal focusing. The major peaks were allowed t o go off-scale, as some of t h e minor peaks were of more interest. Changes in minor constituents from one batch of beer to another were quite apparent when this device was used. Design A performed well in this work as the compounds of interest were low boiling and thus desorbed well a t t h e temperature limit (190 " C ) of design A. T h e upper temperature limit in this design was imposed by t h e presence of a Teflon seat. T h e Vespel seal in design B, however, allowed t h e desorption of higher boiling compounds because of its higher temperature stability (260 "C). Figure 4 shows a cartridge injection of organic volatile compounds found in wastewater using design B. It was necessary t o use the thermal focusing technique with this injection. T h e large sorption cartridge used in design B requires longer desorption times and makes t h e use of t h e thermal focusing technique necessary. If thermal focusing is not used with t h e design B injection system, t h e peaks are broad and often unresolved.
LITERATURE CITED (1) S. J. Faust and J. V. Hunter, "Organic Compounds in Aquatic Environments", Marcel Dekker, New York, 1976. (2) L. H. Keith, "Identification and Analysis of Organic Pollutants in Water", Ann Arbor Science, Ann Arbor, Mich., 1976. (3) R. L. Grob, "ChromatographicAnalysis of the Environment", Marcel Dekker, New York, 1975. (4) R. D. Kleopfer and B. J. Fairless, Environ. Sci. Techno/., 6 (1972). ( 5 ) K. Grob and F. Zurcher, J . Chromatogr., 117, 285 (1976). (6) P. Wedgewood and R. L. Cooper, Analyst (London),81, 42 (1956). (7) L. Keith, "The Analysis of Organic Pollutants in Water and Waste Water", Ann Arbor Science Publishers, Ann Arbor, Mich., 1973, p 30. (8) M. Novotny, M. L. Lee, and K. D. Battle. Chromatographia,7, 333 (1974). (9) W. E. May, S. W. Chester, S. P. Cram, B. H. Gump, H. S. Hertz, D. P. Enagonio, and S. M. Dyszel, J . Chromatogr. Sci., 13, 535 (1975). (10) A . Zlatkis, H. A. Lichtenstein, and A. Tishbee, Chromatographia, 6, 67 (1973). (1 1) B. J. Dowty, L. E. Green, and J. L. Laseter, Anal. Chem.,48, 946 (1976). (12) A. A. Micholson, Otto Meresz, and Brenda Lenyk, Anal. Chem.,49, 815 (1977). (13) A. Zlatkis, U.S. Pa!. Appl. 450500, 12 March 1974, 44 pp. (14) K. E. Rasmussen. J . Chromatogr. Sci., 14, 93 (1974). (15) S. B. Dale, J . Chromatogr. Sci., 7, 389 (1969). (16) J. M. H. Daemen, W. Dankelman, and M. E. Hendriks, J . Chrornatogr. Sci., 13, 79 (1975). (17) B. Versino, M. de Groot, and F. Greiss, Chromatographia, 7, 302 (1974). (18) B. Versina, H. Knoppel, M. de Groot, A. Peil, J. Poelman, H. Schavenburg. H. Vissers, and F. Geiss, J . Chromatogr.. 122, 373 (1976). (19) Harry S. Hertz, Willie E.May, Stephen N. Chesler, and Barry H. Gump, Enwron. Sci. Techno/., 10, 900 (1976).
RECEIVED for review April 20, 1978. Accepted September 8, 1978. P a r t s of this investigation were supported by t h e Air Force Office of Scientific Research, Air Force Systems Command, under contract F44620-76-C-0031.
