Colorimetric estimation of phenols and tyrosine - Analytical Chemistry

Publication Date: November 1975. ACS Legacy Archive. Cite this:Anal. Chem. 47, 13, 2293-2296. Note: In lieu of an abstract, this is the article's firs...
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620nm

644 nm

90

0,75

03

t

A

0,25

20

40

60

80

400

ng/cm3

Figure 2. Standard working curves for beryllium as Chromeazurol S complex at 6 1 1 and 6 2 0 nm. Conditions similar to Figure 1

beryllium concentration. The change is only small a t higher beryllium concentrations, but becomes quite appreciable a t low concentrations. In addition, there is also a slight nonlinearity when absorbance values a t the absorbance maximum are plotted vs. beryllium concentration. As a result, the absorbances of a series of standards measured a t the wavelength of the absorbance maximum of the standard with the highest beryllium concentration (620 nm) will not produce a straight line through the origin when plotted vs. beryllium concentration (Figure 2). Beer’s law, therefore, is not valid for the whole concentration range. The divergence is mainly in the lowest concentration range where the

line shows a bend. This can easily be overlooked, especially when the analyst assumes that his lowest point on the standard working curve may be faulty because of experimental factors when it is not perfectly on the line. Measurement at 611 nm, a wavelength arbitrarily chosen by inspection of the absorbance curves on Figure 1, produces a considerably better straight line going through the origin, though a small divergence is still present for the lowest beryllium concentration measured. A bend in the standard working curve similar to that shown for the measurements a t 620 nm on Figure 2 was also found by us when determining tin spectrophotometrically as its complex with pyrocatechol in the presence of cetyltrimethylammonium bromide in tartaric acid medium, adapting the method described by Dagnall et al. ( 2 ) and H. B. Corbin ( 5 ) . Furthermore, Sommer et al. (3) also show a bent Beer’s plot for beryllium determined with Chromeazurol S using polyvinyl alcohol as micelle-forming agent and triethanolamine as buffer. We therefore suspect that changes in bathochromic shift with changing metal ion concentration similar to that described on Figure 1 may be fairly general when using micelle-forming agents to obtain increased sensitivities during spectrophotometric determination of metals. This has to be taken into account when attempting accurate analytical determinations. In addition, other electrolytes present also can cause a slight change in the bathochromic shift, and the intensity of the color of the beryllium-Chromeazurol S complex in the presence of BDHA changes quite appreciably with foreign electrolyte concentration. I t also is dependent on the kind of electrolyte. Using exactly the same conditions as given for Figure 1,but omitting the 0.1M ammonium chloride will decrease the adsorbance for 100 nanograms of beryllium a t the adsorption maximum by about 15%.Experimental conditions, therefore, should be carefully controlled and reagents used for preparing the standard working curve should match those in the samples when micelleforming reagents are employed for obtaining increased sensitivities in the spectrophotometric determination of metals.

LITERATURE CITED (1) (2) (3) (4) (5)

V. Swoboda and V. Chromy, Talanta, 12, 431 (1965). R. M. Dagnall, T. S. West, and P. Young, Analyst, 92, 27 (1967). L. Sommer and V. Kuban, Anal. Chim. Acta, 44, 333 (1969). H. Nishida, BunsekiKagaku (Jpn Anai.), 20, 1080 (1971). H. B. Corbin, Anal. Chem., 45, 534 (1973).

RECEIVEDfor review April 18, 1975. Accepted August 1, 1975.

Colorimetric Estimation of Phenols and Tyrosine Joseph Chrastil School of Medicine, Vanderbilt University, Nashville, Tenn.

37232

The most commonly used method for the colorimetric estimation of phenols is the color reaction with aminoantipyrine (1-4). This method, based on the oxidation by potassium ferricyanate, is fast and accurate but many ring compounds give the test, and many aromatic hydroxy compounds, especially with an occupied para position, do not react. Several other nonspecific methods are based mostly on coupling of the estimated phenols with diazonium salts. The diazonium salts of sulfanilic acid ( 5 ) , p-nitroaniline

(6-8), m-nitroaniline (9),1-nitroso-2-naphthol (IO), and hydrazinobenzene sulfonic acid ( 1 1 ) were used. The color reactions of phenols with FeC13 (12, 13),HNOz (14, 15), titanium sulfate (16), Millon’s reagent (17) and others have also been used. Several methods have been described for the colorimetric estimation of tyrosine. Millon’s mercury reagent (1820), Folin-Ciocalteau phenol reagent (21-23) or 1-nitroso2-naphthol were used for the estimation of tyrosine (24-

ANALYTICAL CHEMISTRY, VOL. 47, NO. 13, NOVEMBER 1975

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Table I. Colorimetric Estimation of Tyrosine in Mixtures with Amino Acids I W V

z a m a

0

ln

m

a

50

I(

TIME IN MINUTES Figure 1. Influence of H 2 S 0 4 concentration on the color deveiop-

ment of the reaction with tyrosine 1 prnollrnl H20 of tyrosine standard solution was used and analyzed as in the method using 25% (w/w) H2S04 (-), 50% (w/w) H2S04 (- - - -), and 90%

(w/w) HpSO4 (--)

31). The color formed after the reaction of l-nitroso-2naphthol with tyrosine is very unstable and must be decomposed to a more stable yellow complex which must then be separated from the excess reagent by extraction. The whole procedure is accompanied by a loss of the tyrosine caused by nitration and extraction. Although this method is more specific than the two previous methods, it sometimes lacks specificity when applied to plasma or other tissue. The approved fluorometric modification of this method (32) is sensitive to nitration, extraction, pH, and incubation temperature. The color of ceric ammonium nitrate in nitric acid or ammonium hydroxide changes from yellow to orange or red in the presence of alcohols, phenols, and some aromatic amines (33-37). Because of the unstability of this colored complex, several methods, including extrapolation to zero time (36), were proposed to avoid this difficulty. We have found that soluble ceric salts in the presence of hydroxylamine form a stable orange colored complex with aromatic hydroxy compounds (aliphatic alcohols do not react under these conditions). This colored complex obeys Lambert-Beer’s law. The blank is colorless. The reaction is specific for aromatic hydroxy compounds. I t can also be used for the estimation of phenolic groups or tyrosine in biochemical mixtures or protein hydrolysates.

EXPERIMENTAL Estimation of an Aromatic Hydroxy Compound. Two milliliters of a solution containing 10-1000 nmol of an aromatic hydroxy compound were mixed with 0.2 ml of 30% (w/v) hydroxylamine hydrochloride (99% from Sigma) in distilled water. Then 1.0 ml of 50% (w/w) HzS04 (reagent from Fisher Company) was added and mixed. After 1 minute, 1.0 ml of 5% (w/v) (NH4)2Ce(NO& (99.9% from Research Chemical Corp.) in water was added and mixed thoroughly. The colored complex of the cerium and aromatic hydroxy compound was read after 60 minutes a t room temperature vs. the reaction blank on a Beckman Acta V spectrophotometer. Most of these colored complexes absorbed between 320-450 nm depending on the aromatic hydroxy compound. The color intensity was usually linear with the concentration a t 380 nm. The standard curve must be estimated in each case before analysis. The solution of ceric ammonium nitrate in water must be prepared fresh and filtered before use. The specificity of the method was estimated with different compounds dissolved in H20 or dioxane (200 kg/ml). Estimation of Tyrosine in Mixtures with Other Amino Acids or in Protein Hydrolysates. Two milliliters of a solution of tyrosine (tissue homogenate, blood, protein hydrolysate, etc.) were mixed with a 1.0 ml of 15% (w/v) trichloroacetic acid (99.7% from Fisher Company) and centrifuged. Then 2.0 ml of the supernatant 2294

M,xture

Tbrosine content

I I1

5 .OO% 10 .OO%

Tyrosine found

5.08% 9.94% The tyrosine in the mixture with other aminoacids was analyzed as described in the method. Twenty mg of a mixture of amino acids were dissolved in 2.0 ml of 2M H2S04 and used for the method. The mixtures were chosen as an example simulating an average protein amino acid composition. The composition of the mixtures (w/w with 0.00570 accuracy) was as follows: I. alanine 1070,arginine 570, asparagine 1070, cysteine 270, glutamine lo%, glycine 1070, histidine 5%, isoleucine 5%, leucine 10’70, lysine 5%, methionine 2%, phenylalanine 5%, proline 570, threonine 570, tryptophan 1%, tyrosine 570, valine 5%. 11. alanine 1070, arginine 2%, aspaqagine lo%, cysteine 2%, glutamine 1070,glycine lo%, histidine 2%, isoleucine 5%, leucine lo%, lysine 5%, methionine 2%, phenylalanine 5%, proline 570, threonine 570, tryptophan 1%,tyrosine 1070,valine 570. The recovery of tyrosine in an acid hydrolysate of proteins depends on the conditions of the reaction and is usually 87-95%. Using the method for the estimation of tyrosine as described above, our recovery of tyrosine (over 90%) in an acid hydrolysate of bovine serum albumin (from Sigma Chemical Company) was similar to, for example, Hirs, Stein, and Moore (381, who used chromatographic methods. were transferred into clean tubes containing 0.2 ml of 30% hydroxylamine hydrochloride solution in H20. Next 1.0 ml of 50% H2S04 was added and mixed. After 1 minute, 1.0 ml of 5% (NH4)zCe(NO& was added and mixed thoroughly. After 60 minutes a t room temperature, the orange color was read a t 480 nm vs. the reaction blank. The interference by foreign compounds was estimated with a standard solution of tyrosine (1 pmol/ml) in the presence of different compounds (200 pg/ml). A standard curve was constructed for different amounts of tyrosine in H20. The absorbance was linear with the concentration of tyrosine up to 800 nmol/ml. Experiments with different concentrations of H2S04 were performed with the standard solution of tyrosine (1kmol/ml) (Figure 1).

RESULTS AND DISCUSSION Ceric ammonium nitrate can be replaced by other ceric salts (for example sulfate), but the nitrate has several advantages. It is soluble in water (sulfate needs sulfuric acid as a solvent), the color complex with phenolic compound and hydroxylamine is deeper, and the reaction is faster than with ceric sulfate. Cerous salts do not react. Hydroxylamine cannot be replaced by other reagents such as Na2S03, NaN02, Na2S204, or Na2S. The color is completely developed in 30-60 minutes and is stable for at least 24 hours. Optimum conditions were obtained with 50% H2S04 in most cases. The following compounds do not react and do not disturb the color reaction of cerium with aromatic hydroxy compounds in the presence of hydroxylamine: ATP, acetaldehyde, acetic acid, acetoacetic acid, acetone, acetylsalicylic acid, acetylacetone, acetylcholin, N-acetylcysteine, Gacid, aconitic acid, acridine, adenine, adonitol, alanine, albumin, alloxane, 2-amino-5-chlorobenzoxazole, 4-aminoantipyrine, aminolevulinic acid, aminopyrine, amphetamine, n -amyl alcohol, amytal, aniline, p -anisaldehyde, p -anisic acid, p -anisidine, anisole, anthraquinone, 2-anthraquinone sulfonic acid, anthrone, arabinose, arabitol, arachidic acid, arginine, ascorbic acid, asparagine, aspartic acid, benzaldehyde, benzidine, benzimidazole, benzofuran, benzothiazole, benzphetamine, betaine, biotin, n-butyl alcohol, cadaverine, caffeine, camphor, catalase, cephalin, cetyl alcohol, cholesterol, cholic acid, cholin, citruline, codeine, 2,4,6-collidine, corticosterone, cortisone, coumarin, creatine, crotonic acid, 1,3-cyclohexanediol, cyclamic acid, cyclohexane, 1,2-cyclohexanedion, cyclohexanol, cyclohexanone, cyclohexylamine, cysteine, cystine, cytidine, cytochrome c, decyl

ANALYTICAL CHEMISTRY, VOL. 47, NO. 13, NOVEMBER 1975

Table 11. Molar Absorptivities of Some Aromatic Hydroxy Compounds after the Reaction with Ceric Nitrate in the Presence of Hydroxylamine E = (mM cm

-1 )at

Compound

380 nm

450 nm

Tyrosine DOPA Estrone p- Hydroxy phenylpyruvic acid Phenol Resorcinol Hydroquinone Pyrogallol Phloroglucinol p - Aminophenol 0- Aminophenol 1-Naphthol 2-Naphthol p-Hydroxy benzoic acid Guaiacol Orcinol Thymol Eugenol p- Nitr ophenol 0- Nitr ophenol vi-Nitrophenol Salicylic acid Morphine p-Hydroxy acetophenone Vanilline 0-Cresol

1.65 1.60 0 -94 0.76

1.41 0.35 0.19 0.32

1.09 0.94 1.50 1.20 0.79 1.79

0.48 0.33 0.22 0.28 0.25 0.08 0.92 0.32 0.79 0.52 0.63 0.18 0.09 0.76 0.04 0.09 0.15 0.17 0.53 0.03 0.15 0.44

Table 111. Standard Curves of Thymol, p-Hydroxy Benzoic Acid a n d Estronea Thymol Lrmoliml

Absorbance

9-Hydroxy Benzoic Acid poliml

Absorbance

Estrone

umol, ml

Absorbancc

0.06 0.013 0.07 0.038 0.04 0.016 0.30 0.068 0.35 0.201 0.20 0.090 0.60 0.131 0.70 0.406 0.40 0.178 1.20 0.260 1.40 0.820 0.80 0.350 2.40 0.516 2.80 1.601 1.60 0.692 4.80 1.006 3.20 1.384 The compounds were dissolved in dioxane and analyzed as described in the method. The absorbance was measured at 380 nm with a Beckman Acta V Spectrophotometer in cuvettes with 1-cm cell path length.

isoamyl alcohol or ethyl acetate. Tryptophane in high concentrations reacts with a slightly green color with an ab1.73 sorption peak different from tyrosine. 2.45 Except for the estimation of tyrosine in mixtures with 1.13 other amino acids or in protein hydrolysates (Table I), the 1-.99 method can be used for the colorimetric estimation of 0.57 many other aromatic hydroxy compounds. The sensitivity 0.39 of the method depends on the hydroxy compound which is 2.69 analyzed. The extinction coefficients of the colors with 1.04 some aromatic hydroxy compounds are shown in Table 11. 1.74 The colors (which are mostly orange or orange-yellow) usu1.09 ally had wide absorption maximas between 330 and 500 0.83 nm. The intensity of the colors showed linearity with the 1.90 concentration of the corresponding compound at 380 nm, 0.39 which is the convenient wavelength for reading. Some of 1.19 the reactive compounds formed colored complexes which 0.85 absorbed also strongly a t wavelengths close to 500 nm. This could be used for their distinction from other aromatic hyalcohol, dehydroascorbic acid, dehydrofolic acid, diethyldroxy compounds. (For example, tyrosine and DOPA (3,4phthalate, dihydrosphingosine, dihydroxyacetone, dilantin, dihydroxyphenyl-alanine) both absorbed strongly a t 380 N,N-dimethyl-aniline, N,N-dimethyl-p-phenylenedinm, but DOPA did not have a peak a t 480 nm. Or p-aminoamine, dimethylamine, dithioerythritol, ergosterol, erythriphenol did not absorb a t 480 nm, but o-aminophenol did tol, erythromycin, estradiol, estrone, ethanolamine, ethyl absorb, etc.) How the linearity of color intensity is accomalcohol, ethylene glycol, folic acid, formic acid, formaldeplished is shown on examples in Table 111. Different modihyde, furan, fructose, fucose, fumaric acid, geraniol, globufications of the method can be developed for the estimation lin, gluconic acid, glucosamine, glucose, glucuronic acid, of aromatic hydroxy compounds or their mixtures. glutamic acid, glutamine, glutathione, glyceraldehyde, glyceric acid, glycerin, glycine, glycolaldehyde, glyoxal, guaACKNOWLEDGMENT nine, hexobarbital, n-hexyl alcohol, histidine, hydrazine, The technical assistance of C. McKenzie and secretarial hydroxyproline, indazole, indene, 3-indole-glyoxyamide, assistance of P. Chu are gratefully acknowledged. inositol, @-ionone,isoamyl alcohol, isoleucine, isoprene, isopropyl alcohol, ketoglutaric acid, leucine, limonen, linoleic LITERATURE CITED acid, lysine, malic acid, methionine, methyl alcohol, methyl (1)E. Emerson, J. Org. Chem., 8, 1692 (1943). ethyl ketone, N-methylaniline, NADPH, nicotinamide, ni(2) S.Gottlieb and P. E. Marsh, lnd. Eng. Chem., Anal. Ed., 18, 16 (1946). trobenzene, norvaline, n-octyl alcohol, octylamine, oleic (3) E. F. Mohler and L. N. Jacobs, Anal. Chem., 29, 1369 (1957). (4)R. J. Lacoste, S. H. Venable, and J. C. Stone, Anal. Chem., 31, 1246 acid, ornithine, oxalacetic acid, oxalic acid, oxytocine, pan(1959). tothenic acid, perchloric acid, phenylalanine, phenylpyru(5)J. J. Fox and A. J. H. Gange. J. SOC. Chem. lnd., 39, 260 (1920). vic acid, phosphoric acid, phytol, prephenic acid, progester(6)J. Moir, J. S. Afr. Chem. lnst., 5 , 8 (1922). (7)R. C. Theis and R. S. Benedict, J. Biol. Chem., 61, 67 (1924). one, proline, n -propyl alcohol, purin, pyridine, pyridoxal, (8)C. H. Rayburn, W. R. Harlan, and H. R. Hanmer, Anal. Chem., 25, 1419 pyridoxine, pyrimidine, pyrrole, pyruvic acid, reduced (1953). (9)E. V . Schvitzhoffen, Z. Anal. Chem., 145, 184 (1954). FMN, reduced glutathione, rhamnose, ribose, sarcosine, se(IO)J. Mlodecka, Chem. Anal. (Warsaw),4, 45 (1952). doheptulose, serine, shikimic acid, sphingosine, squalene, (11) C. R. Tallon and R. D. Hepner, Anal. Chem., 30, 1521 (1958). succinic acid, sulfanilamide, testosterone, tetrahydrofolic (12)E. F. Wespand W. R. Brode, J. Am. Chem. SOC.,58, 1037(1934). (13) S.Soloway and S. H. Wilen, Anal. Chem., 24, 970 (1952). acid, thiamine, thioctic acid, thioglycolic acid, thiophene, (14)L. Nicolas and R. Burel, Chim. Anal., 38, 316 (1956). thiophenol, thiourea, thyroxine, threonine, thymine, tocof(15)N. D. Cheronis, J. E. Entrikin, and E. M. Hodnett, "Semimicro Quantitative Analysis", John Wiley & Son, New York, 1965,pp 406-410. erol, trichloroacetic acid, UDPGA, ubiquinone, uracil, urea, (16)P. H. Caulfield and R. J. Roblnson, Anal. Chem., 25, 982 (1953). uric acid, urotropin, valine, and xanthine. Diphenylamine, (17)M. T. Koks, Pharm. Weekblad, 68, 557 (1931). anthranilic acid, indole, and some indole derivatives also (18)F. W. Bernhart and R. W. Schneider, Am. J. Med. Sci., 205, 636 (1943). (19) S.F. Velick and E. Ronzoni. J. Biol. Chem., 173, 627 (1948). form colored products. Extraction procedures can be used (20)D. Y. Hsia. "Inborn Errors of Metabolism", Year Book Publ.. Chicaao, for the quantitative separation of these products from the Il ., 1959,p 306. (21)0.Folin and V. Ciocalteau, J. Biol. Chem., 73, 627 (1927). phenol-cerium complexes. The color of indole derivatives, (22)F. R. Jevons, Biochem. J.. 89, 621 (1963). for example, can be quantitatively extracted from the cer(23)R. A. McAllister, J. Med. Lab. Techno/.,28, l(1969). ium colored complexes of aromatic hydroxy compounds by (24)0.Gerngross. K. Voss, and H. Herfeld. Ber., 66B, 435 (1933). 2.29

ANALYTICAL CHEMISTRY, VOL. 47, NO. 13, NOVEMBER 1975

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(25) (26) (27) (28) (29) (30) (31) (32) (33) (34)

L. E. Thomas, Arch. Biochem., 5, 175 (1944). J. Giral, An. lnst. Invest. Clent. Univ. NuevoLBon, 1, 115 (1944). S. Udenfriend and J. R. Cooper, J. Bial. Chem., 106, 227 (1952). D. B. McCornick, Anal. Siochem., 37, 215 (1970). K. Uehara, J. Biochem. (Tokyo),68, 119 (1970). R. Hakanson, Anal. Blochem., 51, 523 (1973). J. A. Ambrose, J. Clin. Chem., 20, 505 (1974). W. K. Wong, M. E. Flynn, and T. Inouye, Clln. Chem., I O , 1098 (1984). F. M. Schemjakin, 2.Anorg. Allgem. Chem., 217, 272 (1934). F. R. Duke and G. F. Smith, Ind. fng. Chem., Anal. Ed., 12, 201 (1940); 17, 572 (1945).

(35) (36) (37) (38)

V. W. Reid and R. K. Truelove, Analyst, 77, 325 (1952). V. W. Reid and D. G. Salmon, Analyst, 80, 704 (1955). V. Kratochvil and SobBslavsky, Chem. PrCm., 6, 515 (1956). C. H. W. Hirs, W. H. Stein, and S. Moore, J. Bid. Chem., 211, 941 (1954).

RECEIVED for review June 20, 1975. Accepted August 4, 1975. This work was by grants from the NIH

(GM-15431, GM-21949).

Removal of Trace Elemental Impurities from Polyethylene by Nitric Acid R. W. Karin,’ J. A. Buono,2 and J. L. Fasching3 Department of Chemistry, University of Rhode Island, Kingston, R.I. 0288 7

The importance of trace-element techniques can best be emphasized through the increasing list of applications and developing instrumentation ( I , 2 ) . With this added sophistication, it becomes mandatory to document and reduce all levels of contamination as much as possible (3). Not only do reagents contribute, but all materials that come into contact directly or indirectly with standards and samples must be considered possible sources of contamination. The contamination problem can be best controlled by the use of inert apparatus whenever possible ( 4 ) . Because of inertness and relative cleanliness, polyethylene, Teflon, and high-purity synthetic quartz have been selected as the best container materials ( I , 3). When involved with large sample populations, polyethylene is principally used instead of Teflon and quartz because of economic considerations. A large number of scientists have employed a wide range of techniques for preparing materials for use in trace analysis ( I , 3-11). From these studies, only a few documented their cleaning procedures with limited results. We report in this article some additional studies on the subject. In this work, the use of nitric acid for leaching polyethylene as a cleaning step in preparation for trace element analysis was investigated. Reagent grade (concentrated) and 8N nitric acid were used to evaluate the usefulness of leaching trace impurities from polyethylene. The leaching process was studied for a period of four days to determine the optimum cleaning period.

topes being determined, the polyethylene pieces were irradiated upon completion of each 24-hour leaching period to provide the same type of time-related information. The samples used for longlived isotope determinations were irradiated, leached, and then counted a t the end of each 24-hour cleaning cycle. Liquid standards were heat sealed in pre-cleaned polyethylene tubing and were prepared under controlled conditions using a laminar-flow clean-bench. Three standards were used to avoid chemical incompatibility and spectral interferences. One of these contained elements having isotopes with short half-lives and the remaining two were used for elements with isotopes having long halflives. A 0.65% Co-A1 wire was used to monitor the neutron flux distribution during long irradiations. Activation Analysis. All irradiations were carried out a t the Rhode Island Nuclear Science Center. The nuclear reactor is a swimming pool reactor with a 2-megawatt power rating. Pneumatic irradiation tubes were used to provide rapid handling of samples. The reactor neutron flux was 4 X 10l2n/cm2-sec and the thermal-to-fast neutron-flux ratio is 40 in the pneumatic tube location. Ten-minute irradiations were used for short half-life isotope determinations and activation of the long half-live isotopes was performed with a 7-hour irradiation. These two types of irradiations could provide information for 13 elements (Table I). Short-lived isotopes were counted following a decay period of 3 minutes. Samples were counted first and standards were counted immediately upon completion of the first count. Both sample and

Table I. Isotope Information Used in Activation Analysis Short-lived isotopes Isotope

EXPERIMENTAL Sample Preparation. Polyethylene samples were prepared using two different techniques. In one series of experiments, tops from polyethylene snap-top containers were carefully removed without the use of metal tools. They were placed into two large tanks containing 8N and concentrated nitric acid. Samples were systematically leached up to four days. Five pieces were removed from each tank of acid a t the end of a 24-hour cleaning cycle and rinsed with distilled, deionized, and filtered water. Samples were air-dried in a laminar-flow clean-bench to prevent particulate contamination. Heat sealed polyethylene bags were used to contain samples for irradiation. Prior to use, the bags were cleaned by leaching for several days in 8 N nitric acid. In the second series of experiments, pieces of polyethylene tubing were first irradiated and then systematically leached. In the case of the short-lived iso-

56Mn 66cu 24Na 2V 391 28Al

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ANALYTICAL CHEMISTRY, VOL. 47, NO. 13, NOVEMBER 1975

Gamma-ray, keV

846.7 1039.0 1368.6 1434.2 1642.0 1778.9

Long-lived isotopes

S t a n d a r d one

98Au lz2Sb lZ4Sb

64.7 h r 64.3 d a y 60.3 day 40.2 hr

411.8 564 .O 602.7 1596.6

27.8 d a y 35.3 h r 35.3 h r 83.9 d a y 5.3 y r 5.3 y r

320.1 554.3 776.5 889.3 1173.1 1332.5

S t a n d a r d two

51~r Present address, New England Nuclear Corporation, Atomlight Place, North Billerica, Mass. 01862. Present address, AID Division of Fisher Scientific, 590 Lincoln Street, Waltham, Mass. 02154. T o whom correspondence concerning this article should be addressed.

Half-life

2.6 h r 5.1 min 15.0 h r 3.8 min 37.0 min 2.2 min

82Br “Br

%c 6OCO 6OCO