Anal. Chem. 1986, 58,1599-1600
under these chromatographic conditions may then be calculated. Since each sample contains both of these internal standards, chromatographic resolution may be evaluated for each GCIMS TCDD analyses to ensure that quality assurance requirements for chromatographic isomer specificity have not been exceeded. This is also demonstrated in Figure 1. 3. Minimum Level of Detection. Verification of the minimum level of detection (MLD), especially for samples below detection limit (ND), is a complex problem. Reanalysis of every ND after spiking with the analyte a t or just above MLD is inefficient and costly. This method, however, must be used a t least on a subset of NDs to establish credibility for reporting MLD. Verification of MLD for each sample is certainly an essential part of a monitoring study; however, alternative techniques each have positive and negative points to consider. One solution would be to spike each sample with an additional labeled analyte, such as [13C12]2,3,7,8-TCDD. Although this technique would provide an analysis for the “proper” compound, i.e., 2,3,7,8-substituted isomer, one would have to monitor masses other than for natural TCDD. If an alternative natural TCDD were used, correct masses could be used but they must be monitored at a non-2,3,7,8-TCDD GC retention time. We have chosen the second alternative, and spike each sample with natural 1,2,3,4-TCDD at 5.0 times the target MLD and monitor for TCDD masses through an area containing the retention time window for both 1,2,3,4-TCDD and 2,3,7,8-TCDD. We have found this very beneficia1 for establishing that the sample preparation methodology and GC/MS operating parameters will allow for the quantification
1599
of a natural TCDD near MLD that will meet signal to noise ( S I N )and ion ratio quality assurance criteria (3). However, because fish uniquely bioaccumulate only 2,3,7,8-TCDD of all of the 22 TCDD isomers (2), this technique will work for fish (or most other biological tissues), but it may not work for other matrices where all 22 TCDD isomers can potentially be found at high levels. 4. Additional Considerations, Another technique we now use for high-resolution mass spectrometric analysis of 2,3,7,8-TCDD is to monitor the molecular ion of diiodobenzene (mlz 329.8399) as a lock mass ion in place of one of the ions from perfluorokerosene. We have observed an overall reduction in base-line noise for all channels and an increase in S I N of 2-3 times. We believe this is caused by having fewer stray ions in the analyzer, which can create background signals and increase instrumental noise.
Registry No. TCDD, 1746-01-6. LITERATURE CITED (1)
Tlernan, T. 0. In Chlorlnated Dioxlns and Dibenzofurans in the Total Environment; Choudhary, G., Keith, L. H., Rappe, C., Eds.; Butter-
worth: Boston, MA, 1983; Chapter 13. (2) Kuehl, D. W.; Cook, P. M.; Batterman, A. R.; Lothenback, D. B.; Butterworth, B. C.; Johnson, D. L. Chemosphere 1985, 14(5), 427. (3) Analytical Procedures and Quality Assurance Plan : National Dloxln Strategy; U S . Environmental Protection Agency, EPA/600/3-85-0 19, 1985.
RECEIVED for review November 25, 1985. Accepted March 18, 1986.
Micro Vacuum Distillation of Radioactive Liquids Elaine B. Winshell*’ and Robin P. Ertl
Department of Ecology and Evolution, State University of New York at Stony Brook, Stony Brook, New York 11794 A common procedure in the analysis of biochemical pathways is to isolate intermediates synthesized in the cell from radiolabeled precursors. Often, as the final step, small volumes of dilute solutions containing the product must be concentrated or in some cases separated from precursors by vacuum distillation. Special handling is required because of the small sample size and the possible contamination of vacuum lines and traps with hazardous material. This paper describes a self-contained apparatus that facilitates the vacuum distillation of small volumes of potentially hazardous material in an entirely closed system. The apparatus can distill from less than 1-5 mL, depending on the size of the components used. Furthermore, the low cost of one unit, approximately $0.50, as opposed to the expense of miniaturized distillation apparatus, makes it possible to run multiple samples simultaneously. EXPERIMENTAL SECTION Apparatus and Procedure, A 10-mL disposable polypropylene syringe is attached to a 20-gauge needle, 1.5 in. long, bent at an obtuse angle (Figure 1). The plunger of the syringe is drilled across its diameter at 1-in. intervals to accommodate at 1-in. finishing nail. A 1-dram screw-top glass vial fitted with a rubber stopper (a stopper from a Vacutainer brand tube with an outer diameter of 10.25 mm fits well) is filled to no more than one-third its volume with liquid. The syringe is attached via the ‘Present address: Ramapo College of New Jersey, School of Theoretical and Applied Science, Mahwah, N J 07430.
x
w
Flgure 1. Micro vacuum distlllatlon apparatus: a, 20-gauge needle: b, 10-mL syringe; c, finishing nail; d, rubber stopper; e, l-dram vial.
needle through the stopper, and the liquid is brought to temperature in a water bath with the syringe resting on an adjacent ice bath. In order to prevent the stopper from popping as the temperature rises, a small negative pressure is applied. When equilibration has occurred, the full vacuum is applied by pulling the plunger slowly and fasteningit in place, at any desired volume, with the finishing nail passing through it and resting on the flange of the syringe. The syringe is covered with ice, and in a short time the liquid distills and condenses in the barrel. When the distillation is completed, the needle is removed from the stopper, the nail is removed, and the liquid is efficiently transferred to (for example) a scintillation vial through the needle. Reagents and Procedure. This system was employed to measure the differences in glucose flux among individuals that are genetically variable at the phosphoglucose isomerase (EC 5.3.1.9) locus in gills of Mytilus edulis (blue mussel). Weighed gills were incubated in a saline medium ( I ) containing 5 mM glucose and 0.2 pCi/mL ~-[2-~H]ghcose (Amersham). The glucose is phosphorylated in vivo and transported into the cells as glucose 6-phosphate. Phosphoglucose isomerase catalyzes the reversible
0003-2700/86/0358-1599$01.50/00 1986 American Chemical Society
1600
ANALYTICAL CHEMISTRY, VOL. 58, NO. 7, JUNE 1986
isomerization of glucose 6-phosphate to fructose &phosphate via an ene-diol intermediate resulting in the partial detritiation of the second carbon ( 2 , 3 ) .In this system, detritiation also occurs during the tricarboxylic acid cycle and the pentose phosphate shunt. After incubation, the reaction was terminated by adding an equal volume of 6% perchloric acid. By use of the apparatus described, tritiated water was recovered from a 1-mL aliquot of the assay mixture in approximately 10 min at a temperature of 75 "C with a reduced pressure, obtained by increasing the volume of the syringe by 10 cm3.
RESULTS AND DISCUSSION In a typical experiment, the reaction mixture containing was incubated for 2 6718 f 243 dps/mL of ~-[2-~H]glucose h at 29 "C. The tritium released into the reaction mixture was isolated by vacuum distillation as described above yielding a distillate with 725 f 13 dps/mL. A blank, lacking gill tissue, was incubated in parallel and treated in the same way. Background radioactivity due to carryover the precursor as well as distillation of tritiated water contaminating the original material was 123 f 3 dps/mL or, in this case, 1.8% of the added radioactivity. Average background from nine replications was 2.54% of the counts added. Variation in the detritiation of the reaction mixture ranged from 500 to 1167 dps/mL at 29 "C and represented differences among individual animals.
This apparatus can be used for aqueous solutions as described above and may also be used without modification for distillations of most low-molecular-weight hydrocarbons and polar organic solvents because of the chemical resistance of the polypropylene syringe. The synthetic rubber present may swell in the presence of some solvents but should not result in the loss of vacuum or sample.
Registry No. D-Glucose, 50-99-7;D-[2-SH]glucose,22348-49-8; phosphoglucose isomerase, 9001-41-6; tritium, 10028-17-8. LITERATURE CITED (1) Zaba, B. N.; Davles, J. I. Mar. 6loI. Lett. 1980, 1 . 235-243. (2) Katz, J.; Rognstad, R. I n Current Toplcs In CelIukr Reguletlon; Horecker, 6. L., Stadtman, E. R., Eds.; Academic Press: New York,
1976; VOl. IO, pp 237-248. (3) Rose, I. A. PhlIos. Trans. R . Soc. London, 6 1981,293, 131-143.
RECEIVED for review November 18, 1985. Accepted March 3,1986. This work was supported by NIH Grant GM 21133 to R. K. Koehn. It is contribution no. 573 of the Department of Ecology and Evolution at the State University of New York at Stony Brook.
CORRECTION
Equivalent-Circuit Modeling of a Heat-Flux Differential Scanning Calorimetry Cell. Analysis of Thermal Resistance Factors and Comparison with Experimental Data Guang-Way Jang and Krishnan Rajeshwar (Anal. Chern. 1986,58, 416-421). Equation 16 should read dqs _ d9R _ - - - TSH- Ts + TRH- TSH_ TP - TRH+ dt
dt
RS
RD
RD'
TR- Ts RG Equation 20 should read d(dq/dt) -
+ ~ ( T -R Ts) RG!
