factants being analyzed. The surfactant analysis can only be expressed as mgA. as Azure A Active Substances (AAAS).
(IO) L. K. Wang. J. Y. Yang, and M. H. Wang, J. Am. Water Works Assoc., 67(1), 6-8 (1975). (11)J. V . Steveninck and J. C. Riemersma, Anal. Chem.. 38, 1250-1251 (1966). (12)L. K. Wang, "Improved Colorimetric Methods and Field Test Kits for Analyzing Anionic Surfactants in Water and Wastewater," Tec. Rep. No. ND-5296-M-4, Calspan Corporation, Buffalo, NY, pp 1-5 1, August
LITERATURE CITED (1)"Standard Methods for the Examination of Water and Wastewater", 13th ed., American Public Health Association, Washington, DC, 1971,p 339. (2)"Methods for Chemical Analysis of Water and Wastes", U.S. Environmental Protection Agency, Washington, DC, 1974,p 157. (3)L. K . Wang, J. Am. Water Works Assoc., 67 (1).19-21 (1975). (4)L. K . Wang, W. W. Shuster. and P. J. Panzardi, J. Am. Water Works Assoc., 67 (4),182-184 (1975). (5) A. S.Weatherburn, J. Am. OilChem. Soc., 28, 233-235 (1950). (6) G. R . Edwards and M. E. Ginn. Sewage Ind. Wastes, 26, 945-953 (1954). (7) M. E. Turney and D. W. Cannell, J. Am. Oil Chem. Soc., 42, 544-546 (1965). ( 8 ) V. W. Reid, G. F. Longman. and E. Heinerth. Tenside, 4(9). 292-304 (1967). (9) V. W. Reid, G. F. Longman. and E. Heinerth, Tenside, 5(3-4), 90-96 (1968).
1973.
RECEIVEDfor review November 18, 1974. Accepted March 12,1975. This research was supported by the Economic Development Administration of Puerto Rico, and an unrestricted research grant (RPI Project No. IBN 25) from Gruman Aerospace Corporation, Long Island, NY, through the School of Engineering, Rensselaer Polytechnic Institute, Troy, NY. The authors thank Arthur A. Burr, Rensselaer Professor, School of Engineering, RPI, for making this research possible.
Wet Ashing of Some Biological Samples in a Microwave Oven Adel Abu-Samra, J. Steven Morris, and S. R. Koirtyohann Environmental Trace Substances Research Center, University of Missouri, Columbia, MO 6520 1
Problems associated with methods for the destruction of organic matter prior to metals analyses have been the subject of many papers and are probably covered most thoroughly by Gorsuch ( I , 2). Wet ashing methods seem to have gained greatest acceptance among workers interested in trace metals analyses. Disadvantages of wet ashing include the close and constant operator attention which is required, the need for special hoods to handle perchloric acid fumes safely, and the danger of explosion if established procedures are not strictly followed. In certain cases, i.e., neutron activation analysis using short lived isotopes, the elapsed time can also be a limitation. These problems can be minimized if wet ashing is done in an adapted inexpensive commercial microwave oven. Acid mixtures are heated internally by the oscillating electromagnetic field, resulting in very rapid, safe, and efficient ashing. Although microwave heating has been reported as an efficient sample drier ( 3 ) ,this appears to be the first account of a modified microwave system suitable for wet ashing.
Procedure. The exact procedure depends to some extent on sample size which has ranged from a few mg to 1 gram in our experiments. The container selected, the volume of acids added, and the time required for ashing vary but none seem to be critical. Typically, 0.5-gram samples of dried plant or animal tissue are placed in 125-ml Erlenmeyer flasks and 10 ml of the nitric-perchloric acid mixture added to each. The flasks are then placed inside the oven and ashed for a preselected time. Progress of the digestion can be followed visually and it is usually terminated when perchloric acid fumes appear (approximately 200 OC). The manufacturer suggests that the oven should not be operated empty or nearly empty for extended periods because of the risk of damage to the magnetron. Therefore, a beaker containing 100 ml of water is placed inside the cavity whenever the total acid volume is small. Procedures for evacuation and trapping of acid fumes required considerable attention. Evacuation by water aspirator, a small vacuum cleaner, and a 6-inch diameter drum fan (Dayton Model 4C006), all were employed. A trap to remove acid fumes from the exhaust gases prior to venting is normally used. Trap materials have included solid CaO, solid CaC03, liquid NaOH, and a water spray.
EXPERIMENTAL Apparatus. Microwave oven: The oven was purchased in a local department store (Montgomery Ward Signature Microwave Oven) and is similar to many other brands sold for home use. It is rated at 600 watts and has a cavity about 24 X 36 X 39 cm. T o protect the inside of the oven from very corrosive acid fumes, a locally made Plexiglas box was placed inside the cavity. Large pieces of metal should not be placed in the oven, but the small hinges needed for the door and the screws to fasten them have caused no problem, probably because they are located in the corner of the cavity. A 1-inch diameter hole was drilled through the side of the oven and the liner to accommodate a glass exhaust port which is connected to a suitable evacuation system. The modified unit was checked using a Narda microline Model 8200 electromagnetic leakage monitor and found to be free of microwave radiation leakage. Reagents. A 4:l mixture of reagent grade nitric and perchloric acids was used for most ashings. In a few cases, nitric acid and 30% hydrogen peroxide were used.
Table I. Analysis Results of 1577 Bovine Liver a n d 1571 Orchard Leaves by Flame Atomic Absorption after Microwave Oven Wet Ashing 1577 Bovine Liver, * g / g
1 Zn Cu
132 193
2 131 194
3 131 196
4
(av)
(NBS value)
131
(131) (194)
(130 10) (193 i 10)
193
1571 Orchard Leaves, w g l g .___~__
1 Pb
Zn Cu
42 27 11.3
2 45 32 11.8
3 45 21 11.7
4 46 22 11.8
(av)
(NBSvalue)
(44) (26) (11.6)
(45 i 3 ) (25 3) (12 i 1)
ANALYTICALCHEMISTRY, VOL. 47, NO. 8, JULY 1975
*
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~~~
Table 11. Analysis Results of 1577 Bovine Liver a n d 1571 Orchard Leaves by Neutron Activation Analysis Using Microwave Oven Wet Ashing 1577 Bovine Liver, Mq/g
(av) As
0.04
0.04
0.04
0.05
Se co
1.1
1.1
1.0
1.0
0.22
0.05
(0.04) (1.0) (0.22)
(NBS value) (0.05)"
(1.1 f 0.1) (0.18)"
1571 Orchard Leaves, U g l g
(av)
As
8.7 2.0
Cr Se Ni
0.09 1.4 Uncertified
9.3 0.06
2.7 0.09
0.07
1.3
RESULTS AND DISCUSSION The microwave oven is a very safe, rapid, and convenient way to wet ash samples. Small samples (0.5 gram or less) can be digested (brought into homogeneous or nearly homogeneous solution) in about 1 minute and completely ashed with fuming perchloric acid in about 3 minutes. With the present low power unit, the time required depends on oven loading. Twelve 0.5-gram samples required 15 minutes for complete ashing. The timing is not critical and in several cases the flasks were taken to virtual dryness whereupon clear, colorless, water soluble inorganic perchlorate crystals formed. Erlenmeyer flasks or test tubes worked better than beakers for ashing. This is apparently due to the increased refluxing action of the H N 0 3 in the flasks. This presents the organic material with a better oxidizing atmosphere. Although a certain amount of refluxing seems advantageous, and, therefore, these two it also prolongs removal of "03 opposing factors must be balanced to achieve optimum elapsed times. The amount of acid mixture added must be adequate for the sample size being digested. In some experiments, the amount of acid was intentionally reduced and the expected vigorous reaction between un-oxidized sample and hot perchloric acid was observed. This usually resulted in a flash inside the container with the production of a considerable amount of carbon residue. The important point learned from these controlled "failures" was that on the samples tested there were no explosions. Seven ml of the acid mixture appeared to be the minimum amount for a 0.5-gram sample and 10 ml provided an adequate safety margin without prolonging the ashing time excessively. No foaming, frothing, or bumping has occurred for any of the samples ashed in the oven to date. Micro ashing can be done by simply scaling down the above procedure. For some analytical techniques, however, the residual perchloric acid can be troublesome. Carbon furnace atomic absorption is one such method. Samples of hair, fingernail, and Drosophila weighing 1-5 mg were treated with 100 p1 of HN03 and placed in the oven for 10 minutes. The samples were then removed, 100 p1 of 30% H202 added, and returned to the oven for 15 minutes. This produced a digest which formed a homogeneous solution and was sufficiently free of organic matter for furnace atomic absorption. Samples of NBS Standard Reference Materials 1571 (Orchard Leaves) and 1577 (Bovine Liver) were ashed and several elements determined by atomic absorption. The results, which are presented in Table I, agree quite closely 1476
ANALYTICAL CHEMISTRY, VOL. 47, NO. 8, JULY 1975
0.06
0.08
1.3
(NBS value)
*
(9.0) (2.4) (0.08)
(2.3)Q
(1.3)
(1.3
(11
(0.08 i-
3)
*
0.01) 0.2)
with the certified values. Neutron activation analysis was used for the determination of As, Se, Co, Cr, and Ni in the same reference materials. In each case, a radiochemical separation was used after post-irradiation digestion of the sample in the microwave oven. The results agree quite closely with certified values (Table 11). Tests are currently under way to determine if losses of trace elements occur. However, it seems unlikely that the behavior will differ significantly from conventional wet ashing. The acid fumes must be efficiently removed from the oven if excessive corrosion is to be prevented. One attempt was made to force air into the oven, thereby sweeping the vapors out. This was unsatisfactory because the slight positive pressure within the liner caused excessive leakage of acid fumes in spite of a rather tight-fitting door. Evacuation by a water aspirator did not provide adequate flow to ensure freedom from leaks and rapid clearing of the cavity a t the end of an ashing. A small vacuum cleaner gave adequate evacuation but was soon attacked by acid fumes which passed through the trap. The most satisfactory means of venting was with a 6-inch diameter drum fan. There is still some attack by acid fumes, but the impellers last for several months and are easily replaced. Trapping of acid fumes is desirable to protect the fan, and the hood into which it is vented. Also, in the case of neutron activation analysis, prevention of escape of radionuclides such as 38Cl and s2Br is very desirable. Solid and liquid bases have been tried but were not very satisfactory because of the rather large air flow with the drum fan and because the filters tended to clog. The best trapping method used thus far is a spray fume scrubber constructed locally and shown schematically in Figure 1. The bulk of the acid fumes goes to the drain and the remainder is safely vented to a standard fume hood. The primary advantage of microwave oven wet ashing is the speed with which the digestions can be carried out. Ashings which normally required 1-2 hours with nearly exhaust
exhaust from oven
drain
Figure 1. Acid fume scrubber
constant operator attention can be done in 5-15 minutes with little or no attention. The time saving will be important in many applications but with neutron activation analysis using short lived isotopes, it is especially valuable. Additional advantages of the method include: 1) It is safe. Bumping and frothing are virtually eliminated and the samples are enclosed if an accident should occur. 2) The need for special perchloric acid hoods is eliminated if the acid fumes are trapped. 3) No special glassware is needed. 4) The air intake to the cavity liner can easily be filtered, greatly reducing the risk of air-borne contamination.
This method of ashing has been in routine, nearly daily use in our laboratory for about six months. Many different types of samples have been successfully digested although data are given here for only a few.
LITERATURE CITED (1) T. T. Gorsuch. Analyst, 84, 135 (1959). (2) T. T. Gorsuch, "The Destruction of Organic Matter". Pergarnon Press, New York, NY, 1970. (3) J. A. Hasek and R. C. Wilson, Anal. Chem., 46, 1160 (1974).
RECEIVEDfor review January 15, 1975. Accepted March 12, 1975.
Determination of Combined Vinyl Acetate in Vinyl Acetate-Vinyl Chloride Copolymers by an Isotope Dilution-Derivative Method D. R. Campbell Research and Development Division, The General Tire & Rubber Company, Akron, OH 44329
The determination of combined vinyl acetate in copolymers with vinyl chloride is an important analysis because of the many advantageous processing characteristics conferred by only minor amounts of the ester. Maintaining the concentration of vinyl acetate within narrow limits permits the synthesis of resins whose properties can be modified to meet stringent end-use requirements. Although relatively rapid analytical methods are desirable for production control, the suitability of such procedures should preferably be established by comparison with a reference method of demonstrated accuracy. The micro oxygen-flask combustion technique for chlorine ( 1 , 2 )is rapid and precise but represents an indirect approach of limited usefulness a t vinyl acetate contents less than approximately 5% and cannot be used for determining vinyl acetate in the important terpolymers that also contain vinyl alcohol. Direct chemical methods, which involve hydrolysis and subsequent determination of the acetate, have been complicated by concomitant dehydrochlorination of the copolymers by alkali. Distillation and titration of the acetic acid after adding sulfuric acid and silver sulfate to the alkaline suspension was first proposed (3, 4 ) , but high results have been reported ( 5 )with this technique. The procedure based on passing the alkaline hydrolyzate over an anion exchange resin in the hydrogen form ( 5 ) requires titration of the eluate for both acid and chloride and has also been reported to yield high results (6). An alternative approach is direct titration of the alkaline hydrolyzate successively with standard acid and silver nitrate solutions ( 7 ) . Conductometric titration with standard sodium hydroxide solution after adding an excess of hydrochloric acid to the hydrolyzate (6) has been employed, as has potentiometric titration of the mixture of acetate and excess alkali used for hydrolysis (8-10). Although these instrumental methods require only a single titrant, the need to establish two end points is a common drawback affecting precision. Infrared methods based on measurement of the absolute absorbance of the carbonyl band in solution (11) or the ratio of carbonyl absorbance to that of carbon-hydrogen absorbance in cast films (12, 13) have been reported, and the potassium bromide pellet technique has also been employed, both with ( 1 4 ) and without (13) an internal stan-
dard. Although copolymers of known vinyl acetate content are desirable for calibration of these infrared procedures, the ester content of polymers employed as standards is usually obtained indirectly from chlorine analysis. The determination of combined vinyl acetate has thus remained of considerable interest, as evidenced by the more recent application of high resolution nuclear magnetic resonance (15)and gas chromatography ( 1 6 )to the problem. This report describes an isotope dilution-derivative method for the direct determination of combined vinyl acetate in copolymers with vinyl chloride that can be used to evaluate or standardize all other methods. A known amount of acetic-2-l4C acid is added to a solution or suspension of the resin in methyl ethyl ketone and the ester is hydrolyzed with sodium hydroxide. The major portion of the sodium acetate-14C is isolated and converted to 2methylbenzimidazole-methyl-14C by means of the Phillips reaction ( 17 ) with o-phenylene diamine,
H
Methyl-labeled acetic acid is used to avoid the kinetic isotope effect that would be expected with the reagent labeled in the carbonyl group.
EXPERIMENTAL Reagents. An aqueous solution of acetic-2J4C acid containing 0.5-1.0 millimole per gram is prepared from either the acid or the anhydride. In this work, the anhydride was used and its specific activity was determined by treating a portion with an excess of aniline to form acetanilide-14C, which was crystallized from water and assayed in dioxane-naphthalene scintillator solution. Methyl ethyl ketone (MEK), reagent grade, is stirred with calcium hydride and distilled. Sodium hydroxide, 1 molar, is prepared in 9:l methano1:water. o-Phenylene diamine is crystallized twice from water with the aid of activated charcoal. The scintillator solution is prepared by dissolving naphthalene (200 grams), PPO (2,5-diphenyloxazole) (14.0 grams) and POPOP (1,4-bis-2-[5-phenyloxazolyl]-benzene) (0.06 gram) in scintillation grade dioxane, diluting to 2 liters and adding 200 ml of methanol. ANALYTICALCHEMISTRY, VOL. 47, NO. 8, JULY 1975
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