DISCUSSION Previously there has been no direct and relatively easy way of determining the blank of very pure, low-blank reagents. In the case of silica, the molybdate reagent contains a small quantity of silica, but this cannot be determined directly because it is necessary t o know the silica content of the water being used for the blank determination. Conversely, the silica content of the water cannot be easily determined unless the silica blank of the reagent is known. By the system described, these two unknown factors may be determined simultaneously, either mathematically or graphically, after a calibration curve has been prepared for the reagents involved. This method may be easily extended to include iron determinations for reagent grade chemicals. After the preparation of a calibration curve, the iron concentration of the material being tested may be determined simultaneously with the reagent blank by running only two rapid and direct colorimetric tests. There are three possible cases which may result in the blank determinations. See Figure 3. In case 1, the reagent blank
is positive and adds to the apparent concentration in the determination. The reagent blank may be greater, equal to, or less than the sample content. In case 2, the reagent blank is zero. In case 3, the reagent blank is negative indicating that the reagent has a demand for the substituent being tested. This may result in the determination of free chlorine, oxidizing or reducing agents. After the preparation of a calibration curve, the use of this method will give a simultaneous check on the reagent while running an analysis. This provides a continual check on the reliability of a reagent. ACKNOWLEDGMENT The authors acknowledge J. P. Sickafoose for his assistance during the preparation of this manuscript. RECEIVED for review July 16, 1971. Accepted December 14, 1971.
Improved Tissue Solubilization for Atomic Absorption A n d r e J. J a c k s o n , Leslie M. Michael, a n d H e r b e r t J. Schumacher Department of Encironmental Health, Unicersity of Cincinnati, College of Medicine, Cincinnati, Ohio 45219
ATOMICABSORPTION SPECTROMETRY has proved to be a very useful analytical tool in the study of the trace metal content of animal tissues. Despite its many applications to metal analysis, there are persistent problems such as sample preparation and reproducibility of results. Organic matter present in the sample is a potential source of interference to the aspiration, atomization, and detection of elements in the flame because of nonspecific matrix effects (I). In order t o eliminate this problem the organic material is usually destroyed or disrupted before the sample is analyzed ( 2 ) . Several methods are currently in use, including wet oxidation, dry oxidation, and oxidative fusion (3). I n our laboratory, we have been investigating the use of Soluene-100 (purchased from Packard Scientific), a quaternary ammonium hydroxide tissue solubilizer specifically formulated for use with toluene and xylene based scintillation counting solutions. Employing this product, one can prepare tissue samples for atomic absorption quickly and with minimal handling thus reducing sample loss or possible contamination from excessive sample preparation. Another major advantage of this procedure is that the organic based solubilizer enhances the sensitivity for the metals investigated. Furthermore, the same sample can be used for liquid scintillation counting ( 4 ) as well as gas chromatography ( 5 ) . (1) G. D. Christian and F. J. Feldman, “Atomic Absorption Spectroscopy,” Wiley-Interscience,New York, N.Y., 1970, p 177. (2) T. T. Gorsuch, “The Destruction of Organic Matter,” Pergamon Press. New York. N.Y.. 1970.. D_ 11. (3) Zbid.,pp 19-41. 14) Packard Technical Bulletin. Downers Grove. Ill. 60515. 1970. (5j Joseph MacGee, Veterans Hospital, Cincinnati, Ohio 45220,
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Table I. Comparison of Aqueous and Soluene Sensitivities Sensitivity,* ppm at primary absorbing line (6) Aqueous Soluene method Zn 0.025 0.01 cu 0.10 0.05 Fe 0.20 0.05 Mn 0.01 0.005 absorption. a Concentration that will give a 1
EXPERIMENTAL Reagents. Reagent grade chemicals were used. The Soluene contains a 2 % (WjV) solution of ammonium-lpyrrolidene dithiocarbamate. Procedure. Weigh the tissue sample into a 50-ml volumetric flask. Add 0.5-1.0 ml of Soluene per 100 mg of tissue. Stopper the flask and let it stand at room temperature for 24 hours. Heating to 60 “C will speed solubilization. All tissues investigated gave a clear, homogeneous, and aspiratable solution suitable for atomic absorption. A threeto fourfold dilution of the preparation is made with toluene. The sample was analyzed by the method of additions (6). Similar results were obtained by plotting absorbances of the metal standards in the Soluene matrix against concentration. All determinations were corrected for any reagent metal content by using a reagent blank. Of the metals investigated, only zinc gave a measurable signal. All samples were analyzed on a Perkin-Elmer Model 303 atomic absorption spectrophotometer using instrument settings suggested for organic (6) “Analytical Methods for Atomic Absorption Spectrophotometry,” Perkin-Elmer, Norwalk, Conn., 1971, pp 2-5.
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Table 11. Comparison of Metal Values Obtained Using Soluene and Literature Values for Several Tissues Literature, ppm Amount Level detected, ppm Zn Cu Fe Zn cu Fe Tissue assayed 30.3 6.0 100 30-32 (8) 6.76-12.5 (IO, 11) 30-220 (8) Rat liver 300 mg 11.2 1.7 30 15 (7) 1.85 (11) 40-90 (8) Rat brain 300 mg Rat plasma 0 . 3 ml 0.9 0.5 2.5 1.07 (9) 0.81 (12) 3.0-5.0 (8) Table 111. Comparison of Soluene and Wet Ashing Values for Several Tissues Soluene range, ppm Wet ashing range, ppm Amount Tissue assayed Zn cu Fe Zn cu Fe Rat liver 100 mg 31.3-31 . 8 5.0-5.3 150.G152.0a 26.5-32.5 3.8-7.5 270.0-272.0 Rat brain 100 mg 15 .0-15.5 3.9-4 .O 27.0-28.0 14.3-16.4 2.9-3,6 19.1-33 .O Rabbit plasma 0 . 1 ml 1.2-1.5 0.7-0.8 1.5-2.0 0.8-1.0 0.6-0.9 2.0-3.5 Rat muscle 100 mg 18.0-18.1 2.0-2.1 15.0-15.3 18.0-23 .O 1 . O-1 . 6 10.0-13.0 These values are lower than those obtained for wet ashing, but they are within the range of those cited in the literature.
solvents by the manufacturer (7). The unit is equipped with a recorder read out and Boling burner. RESULTS AND DISCUSSION
The applicability of the procedure for Zn, Cu, Fe, and M n was investigated but may not be limited to these elements. Data obtained using the method of additions followed Beer’s law in the concentration range of 0.05 to 0.50 pg/ml. The same tissue analyzed using standard solutions containing known concentrations of the metals in a Soluene matrix gave values in agreement with those obtained by the method of additions. Flame scattering at a non-absorbing wavelength was approximately 3-5 absorption. (7) “Analytical Methods for Atomic Absorption Spectrophotometry,” Perkin-Elmer, Norwalk, Conn., p 17. (8) W. S . Spector, “Handbook of Biological Data,” W. B. Saunders Company, Philadelphia, Pa., 1956, pp 50-72. (9) E. I. Dreosti, Shyy-Hwa Tao, and L. S . Hurley, Proc. SOC. Exptl. Biol. Med., 128, 169-174 (1968). (10) D. D. Grant and E. J. Underwood, Aust. J . Exptl. Biol., 36, 339-346 (1958). (11) S. La1 and T. L. Sourkes, Biochem. Med., 4, 260-276 (1970). (12) D. H. Cox and D. L. Harris, J . Nutr., 70,514-520 (1968).
Comparative sensitivity between the organic medium and aqueous standards are shown in Table I demonstrating the greater sensitivity of the Soluene method. Table I1 shows that our values are within the range of those reported in the literature. I n a similar manner, Table I11 shows that values obtained using the Soluene method are comparable t o those obtained on the same tissue sample using the accepted wet ashing procedure. Precision as estimated by the coefficient of variation (13) was calculated to be 5.1 %. The method provides a quick, simple, and reproducible procedure for preparing a homogeneous and aspiratable tissue sample. This method is also capable of giving reproducible data with a sample size of 50-100 mg of tissue. Because of the conditions of sample preparation, the sample is likely to be less contaminated and, in addition, one obtains increased sensitivity because of the organic medium.
RECEIVED for review August 2, 1971. Accepted December 14,1971. (13) H. Kaiser and B. Meddings, A t . Absorption Newslert., 6 (2), 28 (1967).
Application of Multiple Internal Reflection Spectrometry to Aircraft Materials Evaluation T.T. Bartels Engineering Laboratories, McDonnell Aircraft Company, St. Louis, Mo. 63166
THE PROPERTIES of many polymeric materials make them suited for use in modern aircraft construction. Polyesters and silicones are used in electrical casting, encapsulating, and potting and sealing applications; urethanes are used in chemical foams t o render fuel tanks inert and as protective coatings; polycarbonates are used in internal stores containers and in canopy applications; polyimides and epoxies are used in resin matrix composites for structural hardware and in adhesives. These applications, of course, represent only a few of the many possible. The variety, complexity, and widespread application of current polymeric materials require rapid test techniques for their characterization and evaluation.
Infrared spectrometry is one of the most convenient single techniques available for determining molecular structure a n d for qualitative identification. Precise quantitative determinations also are possible, provided the components of the material being analyzed possess unique absorption bands separated from absorptions of other components. The difficulty of evaluating many of the previously mentioned polymeric materials by conventional transmission infrared spectrometry lies in the area of sample preparation. Cured materials are generally chemically resistant and thoroughly cross-linked; these properties make solution spectrometry and “mull” techniques tedious and time-consuming. MicroANALYTICAL CHEMISTRY, VOL. 44, NO. 6, M A Y 1972
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