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(44) Harrison, A. G. Chemical Ionization Mass Spectrometry: CRC Press: Boca Raton, FL, 1983; pp 87-127. (45) Middlemiss, N. E.: Harrison, A. G. Can. J . Chem. 1878, 5 7 , 2827-2833. (46) Ryhage, R.: Stenhagen, E. J . Lipid Res. 1980, 7 , 361-390.
RECEIVED for review September 8, 1986. Accepted March 2,
1987. This work was supported by the Midwest Center for Mass Spectrometry, a National Science Foundation Regional Instrumentation Facility (Grant No. CHE-8211164), and by the National Science Foundation (Grant No. CHE-8320388). Presented in part a t the 34th Annual Conference on Mass Spectrometry and Allied Topics, Cincinnati, OH, June 1986.
AIDS FOR ANALYTICAL CHEMISTS Wet Ashing in Biological Samples in a Microwave Oven under Pressure Using Poly(tetrafluoroethy1ene) Vessels Prasad Aysola, Perry Anderson, and Cooper H. Langford*
Department of Biological Sciences and Chemistry, Concordia University, 1455 de Maisonneuve Boulevard West, Montreal, Quebec, Canada H3G l M 8 Gorsuch ( I ) has described the problems associated with the oxidation of organic matter in biological samples. This is a step that must precede trace metal determination by atomic spectroscopy. Treatment with acid on a hot plate typically requires 1-2 h. The use of a microwave oven is an attractive possibility for acceleration of the process (2-5). Koirtyohann e t al. (2) and Barrett e t al. ( 4 ) modified microwave ovens by adding an exhaust port. Nadkarni (3)exploited an unmodified oven by using a Pyrex vacuum desiccator as a pressurizable vessel. They report significant losses of Cu (26%) and P b (20%). Matts e t al. (6) tried polycarbonate pressurizable vessels, but the plastic quickly became opaque and brittle. We have reevaluated the prospects for use of an unmodified microwave oven with pressurized vessels. We found that Pyrex vessels gain heat in the glass quickly. We substituted Teflon TFA brand fluorocarbon resin for polycarbonate and find it has superior chemical and mechanical properties. We now report a 60-s pressure vessel procedure using an unmodified commercial oven.
EXPERIMENTAL SECTION Apparatus. The microwave oven was a 700-W commercial model available locally. The Teflon PFA containers were Savillex Corp. (Minneton, MN) 60-mL vessels 0.11 in. thick in wide-mouth microwave-oven-proofplastic containers. Atomic spectra were recorded with flame AAS employing the following wavelengths: Cu, 324.8 nm; Fe, 248.3 nm; Cd, 228 nm; Ca, 422 nm; Cr, 357.9 nm; Pb, 283.3 nm; Zn, 213.9 nm. The spectrometer was a Perkin-Elmer Model 503 equipped with a 4-in. burner. Materials. Water was double deionized. All acids were Fisher Scientific "trace metal grade". Standards were Fisher Scientific certified grade. The samples were NBS bovine liver (1577) used as is or doped. Procedure. Samples of 0.25 g were placed in Savillex vessels along with 1.5 mL of H2S04and 1.5 mL of HN03. The cap was screwed on tightly with the plastic wrench supplied by Savillex Corp. The sample vessel was then placed in a wide-mouth plastic container which was closed with a screw cap. A small beaker containing 20 mL of water was placed in the oven along with the sample container to avoid damage to the magnitron. Each sample was heated for 60 s at the maximum power setting of the oven. On the basis of the manufacturer's literature, we estimate pressures in the vessels at near 100 psi and temperatures near 200 "C. The container was removed and cooled in an ice bath for 5 min. Then the contents were diluted to 25 mL volume with purified water. Conventionalflame AAS procedures were followed. An acid blank containing the same amount of H2S04and HNOB was used.
Table I. Analysis of NBS Bovine Liver by Flame AAS after Wet Ashing under Pressure in a Microwave Oven"
element
expected concn, pg/g
concn found, pg/g
copper
158 i 7 194 i 20 123 i 8
158 i 3 202 i 8 133 i 2
iron zinc (I
Analyzed values are the mean of three replicates.
Table 11. Recovery of Spiked Cd, Cr, and Pb from NBS 1577 Bovine Liver after Wet Ashing under Pressure in a Microwave Oven"
a
element
amt added, pg
amt recovered, bg
cadmium chromium lead
25 25
26.3 f 0.3 24.4 f 0.4
100
96.3 f 1.9
Analyzed values are the mean of three replicate spikes.
Table 111. Recoveries of Added Nickel in Fish Tissues after Wet Ashing under Pressure in a Microwave Oven"
amt added, pg
amt recovered, pg
10.0 30.0 50.0
9.6 f 0.4 29.5 i 0.8
liver
muscle
10.0
30.0 kidney
10.0
30.0
49.7 f 0.6
9.9 f 0.5 29.9 i 1.1 9.2 f 0.3 29.1 f 0.8
"Analyzed values are the mean of three replicate samples. RESULTS A N D DISCUSSION
Table I reports data for determination of Cu, Fe, and Zn for which NBS values are available. Table I1 displays recovery of spikes of Cd, Cr, and Pb. Throughout recovery is satisfactory. There is no apparent matrix effect but, as is common, some liver samples have residues. The samples with residues are filtered and washed with water and the filtrates are diluted to 25 mL with purified water. Table I11 shows a few results comparing three different types of tissue.
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We have used the microwave oven/pressurized vessel technique for 6 months now. It has been applied to over 100 plant, fish, and soil samples, many of which we have cross reference data from conventional wet ashing procedures. The technique seems quite versatile. I t has also been used for iodine analysis in plasma. Recovery of iodide added (0.5 to 3.0 ng/mL) ranged from 93% to 101%.
ACKNOWLEDGMENT The authors wish to acknowledge J. Bureau, who provided nickel-contaminated fish tissue. Registry NO. CU, 7440-50-8; Fe, 7439-89-6; &, 7440-66-6; Cd, 7440-43-9;Cr, 7440-47-3; Pb, 7439-92-1;Teflon/PFA, 26655-00-5.
LITERATURE CITED (1) Gorsuch, T. T. The Distructlon of Orpnic Mefter; Pergamon: New York, 1970. (2) Abu-Samra, A.; Morris, J. S.; Koirtyohann. S. R. Anal. Chem. 1975. 47, 1475-1477. (3) Nadkarni, R. A. Anal. Chem. 1984, 56, 2233-2237. (4) Barrett. P.; Davidowski. L. J.; Penaro, K. W.; Copeland, T. R. Anal. Chem. 1978, 50. 1021-1023. (5) Fisher, L. B. Anal. Chem. 1986, 58, 261-265. (8) Matthes, S.A.; Farrel, R. F.; Mackie, A. J. Tech. Prog. R e p . - U S . , Bur. Mines 1983, No. 120.
RECEIVED for review October 23, 1986. Accepted February 2, 1987. This research was supported by funds from the Department of Education [FCAR], Quebec.
Packing- Induced Brlttleness in Polyimlde- and Aluminum-Clad Fused-Sllica Capillaries Soon M. Han and Daniel W. Armstrong* Department of Chemistry, Texas Tech University, Lubbock, Texas 79409 Since Golay first used capillaries in chromatography (2,2), they have become virtually indispensable to the field. Although the potential of capillary columns was rapidly realized in gas chromatography (GC) they have only recently made significant in-roads in liquid chromatography (LC) (3-9). In large part, this was due to the lack of appropriate “hardware” (e.g. low dead volume injectors, detectors, connectors, appropriate pumps, etc.) needed for optimal use of capillary LC columns. With the advent of fused-silica capillaries (IO) and the greater availability of proper hardware, “microcolumn LC” is becoming increasingly popular ( I I , 1 2 ) , Indeed, packed capillary columns currently may be the most convenient way to obtain very high plate numbers (?lo6) in LC. Fused-silica tubing now has replaced most other column materials in all forms of capillary chromatography. A variety of fused-silica LC microcolumns have been packed in our laboratory. Some columns became very brittle and essentially impossible to handle without breaking. In some cases, one portion of a packed column would remain flexible while another section would be brittle. In this report the origin and cause of brittleness in polyimide- and aluminum-clad fused-silica packed capillaries is examined.
EXPERIMENTAL SECTION A Shimadzu (Model 5A) liquid chromatograph equipped with a reservoir and a vibrator was use&. Figure 1 shows the packing apparatus. The reservoir was stainless-steel capillary tubing (4.6-mm i.d. X 6 cm long and 4.6-mm i.d. X 12 cm long, Supelco, Houston, TX), taped on the vibrator (Pollenex, Chicago, IL). Fused silica tubings with polyimide coating 250-pm i.d. (350-pm 0.d.) and 50-pm i.d. (160 pm 0.d.) were obtained from Polymicro Technologies (Phoenix, AZ)and Hewlett-Packard (Avondale,PA). Deactivated fused-silica tubing with aluminum-clad coating (250-pm i.d.) was obtained from Quardrex Co. (New Haven, CT). A Teflon frit (5 pm porosity, Alltech, Houston, ?x)was inserted into the cross section of the fused-silica capillary tubing to prevent leaking of the packing material. Five-micrometer spherical pcyclodextrin chiral stationary phase (13),CIS, and C8 (Advanced Separation Technologies, Inc., Whippany, NJ) were used as packing materials and were slurried in HPLC grade methanol or acetonitrile (Fisher Scientific Co., Raleigh, NC). The prepared slurry solution was dispersed for 1 min in an ultrasonic cleaner (Model 321, Branson Co., Shelton, CT) and immediately placed in the reservoir by using a syringe. After the reservoir was connected, packing was started by using a high-pressurepump. The 0003-2700/87/0359-1583$01.50/0
pump was used in the constant-pressuremode. The pressure was increased up to 350 to 470 atm and kept at the maximum pressure for 30 min. Vespel ferrules (Alltech, Houston, TX) were used to connect the fused-silica tubing directly to stainless steel unions. Epoxy was used to connect the 250-pm-i.d. capillary tubing with 50pm4.d. capillary tubing. All electron micrographs were taken with an Hitachi S-570 SEM (scanning electron microscope).
RESULTS AND DISCUSSION Polyimide- and aluminum-clad fused-silica capillaries were divided into four groups according to their recent packing history. The first group consisted of new unpacked tubing. The second group consisted of tubing that had been packed one time. The third group consisted of tubing that had been packed a t half the usual pressure (see Experimental Section). The fourth group consisted of tubing which had been repacked or in which the first packing had been particularly slow or completed in two stages. Electron micrographs (EMS) were taken of the four groups of capillaries (see Figures 2-5). Figure 2 gives an end-on view of a new unpacked polyimide fused-silica capillary. Note the good integrity of the fused silica and the even coating of polyimide. Figure 3 is an EM of the cross section of a “once, high pressure-packed” capillary (polyimide coating). Note that there are a few simple cracks in the fused silica. Analogous results were obtained for aluminum-clad columns. High-pressure slurry packing of these capillaries sometimes produced a few simple cracks. A simple crack did not seem to significantly affect either the flexibility or performance of these columns. Indeed, if it were not for the EMSit would have been difficult to discern that any cracks existed under typical microcolumn conditions. These simple, hard-to-detect primary cracks can result from previous packing, improper handling or cutting, or manufacturing defects. Figures 4 and 5 show EMS of cross sections of polyimide- and aluminum-clad capillaries that contained a simple crack and were then repacked. Note the secondary pattern of cracks that emanate from the larger simple cracks. Also note the irregularity of the aluminum coating in Figure 5. The onset of extreme brittleness in both types of capillaries seems to be related to the presence of secondary cracking of the fused silica. Slurry packing or extended high-pressure use of any fused-silica capillary that contains crack(s) often results in more extensive secondary cracking (Figures 4 and 5). Cur0 1967 American Chemical Society
