ANALYTICAL CHEMISTRY
1650 Table 111. Typical Analyses Influent Fe, N’
Concn. Acid. N
0.2148 0.1074 0.2148 0.2148 0.2148 0.1074 0.0215 0
Sample Taken Acid, meq.
Total meq.a
Recovered Total, rneq.”
0.5784
Fe, rneq.“ IO. 74
28.92
39.66
40.0
Leakage. Fe, 2Va 0.000216
0,289’ 0.3064 0.0922 0.0344 0.5616 0.5474 0,5440
10.74 IO.74 10.74 10.74 5.37 1.074 0
28 92 15.32 4 61 1.72 28.08 27.37 27.20
39.66 26.06 1R,35 12.46 33.44 28.44 27.20
39.6 25.8 15.2 12.5 33.2 28.4 27.2
0 000036 0.003036 < O . 003007 < O . 000007 0.000144 1 0 000007 0
present in the solution, the method has been successfully applied to the determination of copper(I1) and iron(I1) mixtures of high acid content. Presumably i t could be adapted t o other similar situations where the analysis of high concentration of both metal cations and the acid is required.
411 normalities calculated as acid.
ACKNOWLEDGMENT ~~
gible fraction of the influent iron in all cases. .4s would be expected, the magnitude of the leakage was directly proportional t o the acid content of the influent. I n routine analysis this method has consistently given results of greater than 1 part per 1000 accuracy, as shown in Table 111. Greater accuracy may be obtained a t the expense of speed by d e creasing the rate of flow, using resin of smaller particle size, and/or increasing the volume of the water wash. With this increased care accuracy of the order of 3 parts per 1000 is possible. I n addition to the present case where only iron(I1) and acid were
~-
The authors ITish t o express their appreriation to Diana Kenny for her assistance in the experimental phaqeq of this work. LITERATURE CITED
(1) Horner, C., Winger, A , . Bodamer. G., and Kunin, R., Ind. Eng. Chem., 47, 1121 (1955).
(2) Samuelson, O., “Ion Exchangers in .Inalytical Chemistry,”
Wiley, Sew York, 1953. RECEIVED for review May 14, 1954. Accepted June 30, 1955. Presented before the Analytical and Microcheniical Group of the Philadelphia Section A r , i E H i c a s CHEMICAL SOCIETY, April 15, 1955.
Determination of Nicotine, Nornicotine, and Total AI kaloids in Tobacco ROBERT H. CUNDIFF and PETER C. M A R K U N A S R. J.
Reynolds
Tobacco Co., Winston-Salem, N. C.
This study was undertaken to provide the industry with a more rapid procedure for the analysis of total alkaloids, nicotine, and nornicotine-type alkaloids in tobacco. According to the proposed method an alkaline tobacco mixture is extracted with benzene-chloroform solution and aliquots of the extract are titrated with standard perchloric acid. The fact that secondary hase-type alkaloids present in tobacco may be quantitatively acetylated provided a means of determining nornicotine-type alkaloids in the presence of nicotine. Ammonia may be eliminated as an interferant in the proposed scheme of analysis.
ICOTIXE in tobacco products may be determined gravimetrically ab the silicotungstate ( I - 3 , 6 , 8 ) ; volumetrically, after liberation and extraction of the free base ( 7 , 11, 16, 19); spectrophotometrically, after isolation of the alkaloid by steam distillation ( 2 1 ) ; and colorimetrically (IS, 2 2 ) . ?;one of these procedures will distinguish nornicotine from nicotine. Since the literature on the determination of nicotine and related alkaloids is voluminous, the references listed are representative rather than inclusive. Bowen and Barthel ( 5 )and Markwood ( 1 4 ) dehcribed methods for separating nicotine and nornicotine b j reaction of nornicotine n ith nitrous acid. Larson and Haag (12 j proposed a colorimetric procedure for the simultaneous determination of these two alkaloids, and among others Houston (9) has presented a chromatographic technique for separation and determination of nicotine and nornicotine. The silicotungstate methods are time-consuming and tend to give erroneous results when appreciable ammonia or ammonium salts are present ( 2 6 ) . Jeffrey ( I O ) compared several alkaloid methods and found that there was satisfactory correlation of rpsults between the conventional procedures investigated when nicotine was the predominant alkaloid, but noted considerable discrepanry of results when mised alkaloid types of tobacco were
analyzed. The Houston chromatographic technique (9) gives precise results and provides a means of determining both nicotine and nornicotine. However, this procedure is not readily adaptable for the routine analysis of a large number of samples. As it was believed that an extraction and titration procedure would provide the most satisfactory approach to a simplified and rapid analysis, the Garner titration method ( 7 ) appeared to be a suitable st,arting point for the investigation. I n the Garner method an alkaline t,obacco paste is allowed to stand overnight in contact wit,h petroleum ether or a similar type of solvent, an aliquot of the hydrocarbon phase is extracted with standard acid, and the nicotine is determined by titration with standard alkali. Although this method is simple, it, too, is time-consuming and involves an indirect titration. By the proposed procedure it was found po alkaloids from an alkaline tobacco mixture with a benzene-chloroform solution after only a 15-minute extraction period, and t o titrate the alkaloids in an aliquot of the hydrocarbon extract directly wit,h standard perchloric acid in acetic acid. It was found also that nornicotine could be quantitatively acetylated with acetic anhydride, and after acet,ylation, only one half of t’he nornicotine equivalent ivas titrated in the nonaqueous system. This, therefore, provided a means of determining nicotine and nornicotine in the same solution. A number of alkaloids and alkaloid derivatives have been identified in tobacco, Although the principal alkaloids in commercial tobacco are nicot,ine and nornicot,ine, related species and hybrid tobaccos may have correspondingly larger amounts of other alkaloids, especially anabasine. Since alkaloids other than nicotine or nornicotine are present in many tobaccos, it mae thought preferable to designate the tertiary amine alkaloids, nicotine-type alkaloids, and the primary and secondary amine alkaloids, including anabasine, nornicotine-type alkaloids. Hence in this work unless definitely specified, the connotation “nicotine” includes the tertiary amine alkaloids, and “nornicotine” includes the primary and secondary amine alkaloids.
V O L U M E 27, NO. 10, O C T O B E R 1 9 5 5 Table I.
1651
Alkaloid bdded, M g .
Alkaloid Recovered Nicotine Iiornicotine
IiOP
nicotine
hlg.
%;
Mg.
%
50.8 50.8 50.8 101 7
34.6 34.6 17.3 34.6
50.9 50.6 50.4 100.6
100.20 99.61 99.21 98.92
33.5
96.82 100.58 93.64 97.98
Mean
34.8 16.2 33 9
99.48
97.2fi
Table 11.
Recovery of Nicotine and Nornicotine When Added to Tobacco Alkaloid Added, M g . __ Alkaloid Recovered 50 28 28 28
8 8 8 8
Nornicotine 15 1 16 7 16 7 16 i
Nicotine
70
Mg. 50 28 29 28
-
4
21 30 08 65
4 7
99 99 102 99
llean
100 06
h
Iiornicotine' 0' llg. IC 14 16 16 16
9 4 4 5
98 98 98 98
68 20 20 80
98 4 i
-
REAGEhTS, SOLUTIOhS, 4VD 4PPARATUS
Acetic acid, 4.C.S. grade. Acetic anhydride, A.C.S. grade. Barium hydroxide, octahydrate, A.C.S. grade. Celite analytical filter aid, Johns-Rlanville Corp. Perchloric acid, 70 to 72% B.C.S. grade. Potassium acid phthalate, primarv standard grade. Benzene-chloroform solution. Miu 900 ml. of benzene with 100 ml. of chloroform. Barium hydroxide solution. A saturated aqueous solution Crystal violet indicator. Dissolve 1 gram of crystal violet it1 100 ml. of glacial acetic acid. Perchloric acid solution, 0.025*V. Dilute 2.1 ml. of 72% perchloric acid to 1 liter with glacial acetic acid. Standardize the perchloric acid solution against potassium acid phthalate according to the procedure of Seaman and Allen ( 1 7 ) . Precision-Shell titrator with modified calomel and glass electrodes was used for the potentiometric titrations. hlodified calomel electrode. Replace the saturated aqueous potassium chloride in a fiber-type calomel electrode with a methanol solution saturated with potassium chloride. Wrist-action shaker, Rlodel BB, Burrell Corp., Pittsburgh, Pa. PROCEDURE
Accurately weigh 2.5 to 3.5 grams of finely ground tobacco into a 250-ml. glass-stoppered Erlenmeyer flask. Add approximately 1 gram of granular barium hydroxide and 15 ml. of barium hydroxide solution. Swirl the flask until the tobacco is thoroughly wetted, adding more barium hydroxide solution if necessary. Pipet 100 ml. of benzene-chloroform solution into the flask, stopper tightly, and agitate vigorously for 15 minutes using a shaking machine, or for 20 minutes if shaken by hand. Add approximately 2 grams of Celite, swirl flask until the filter aid is well dispersed, allow the two liquid phases to separate, and filter the majority of the hydrocarbon layer through Whatr man No. 2 filter paper into a second flask. Pipet 25-ml. aliquots of the filtrate into each of two 125-ml. Erlenmeyer flasks. Pass a stream of air over the surface of the solution in the first flask for 5 minutes to remove any free ammonia that might be present in the extract. Add 0.5 ml. of acetic anhydride to the second flask. T o each flask, add 1 drop of crystal violet indicator and titrate to a green end point with the 0.025N perchloric acid. If the nornicotine content is found to be as high as 25% of the nicotine content, acetylate another 25-ml. portion of the filtrate, add 25 ml. of acetic acid, and titrate potentiometrically to obtain the equivalence point. Calculations.
% total alkaloids, as nicotine
=
% nornicotine
Xicotine
Sicotine
% nicotine
Recovery of Nicotine and Nornicotine from .4queous Solutions
=
VI X N X 32.45 at. of sample (moisture-free basis)
(2Vz - VI) X N X 32.45 wt. of sample (moisture-free basis)
=
2( Vi - Vz) X h ' X 29.64 wt. of sample (moisture-free basis)
where VI = milliliters of perchloric acid required to neutralize nonacetylated aliquot VZ = milliliters of perchloric acid required to neutralize acetylated aliquot N = normality of perchloric acid solution EXPER1,M ENTA L
Solvents and Extraction. Several solvents were investigated These included petroleum ether, benzene, iso-octane, and chloroform. Benzene proved to be more satisfactory as an extracting solvent than the other hydrocarbons investigated. The indicator end point was sharper in benzene solution, and of the solvents tested only chloroform was more efficient as an alkaloid extractant. Chloroform alone was not investigated further as an extracting solvent because of its high specific gravity. A solvent with specific gravity less than 1 would be much easier to separate from a mixture. A solution of 9 parts of benzene and 1 part of chloroform m-as found to have the advantages of both of these solvents. T o test the efficiency of this benzene-chloroform solution as an alkaloid extractant, known mixtures of nicotine and nornicotine were made alkaline and extracted with benzene-chloroform solution, and the alkaloids were determined as outlined in the preceding method. Table I indicates the recovery of these alkaloids from aqueous solutions. Equally good recovery was obtained when known mixtures of these alkaloids were added to tobacco and analyzed by the described method. This is demonstrated by the data in Table 11. Tobacco samples without added alkaloid were analyzed simultaneously to serve as blanks for these determinations. Acetylation. Treatment of primary and secondary amine8 with acetic anhydride to yield neutral actylation products in nonaqueous systems has been reported (18, 20). Since nornicotine contains both a secondary and tertiary amine group, it was believed that the secondary amine group would acetylate in benzene-chloroform solution. When acetic anhydride was added to a benzene-chloroform solution of nornicotine, allowed to react for several minutes, and titrated with perchloric acid, exactly one half of the equivalent was titrated, whereas the total equivalent was titrated in a nonacetylated solution. This acetylation of nornicotine takes place rapidly a t room temperature. Sicotine, being a tertiary base, is unaffected by the acetic anhydride treatment. The amide formed on acetylation of nornicotine caused a false visual end point in the subsequent titration, so that it was necessary to ascertain the equivalence point potentiometrically. This amide formation also affected the visual end point in tobacco samples where the nornicotine content was as high as 25% of the total alkaloid. Hence all tobacco samples in which the nornicotine content is 25% or more of the total alkaloid content should be titrated potentiometrically after the acetylation Samples in which the nornicotine content is less than 25% of the total alkaloid may be titrated satisfactorily by means of the indicator end point procedure. Potentiometric Titrations. Although suitable titration curves were obtained with the regular glass-calomel electrode system in benzene-chloroform solutions, much more satisfactory curves were obtained if 25 ml. of glacial acetic acid were added to the benzene-chloroform solutions prior to titration, and if the calomel electrode was modified by replacing the aqueous potassium chloride with methanolic potassium chloride. as alkaloid extractants.
ANALYTICAL CHEMISTRY
1652 Table 111. Reproducibility of Extraction Procedure as Determined by Analysis of Flue-Cured and Burley Tobacco Samples % Total 7Z Total Sample Flue cured No. 588
Alkaloids as Nicotine 2,44 2.43 2.43 2 44 2.43 2.40 2.42 2.43 2.44 2.46
Mean Standard deviation
Table IV. Total Alkaloid as Nicotine,
Sample Burley No. 589
2.43 0.016
Alkaloids as Nicotine 4.09 4.10 4.10 4.11 4.09 4.10 4.12 4.11 4.10 4.13
4.11 0.013
Reproducibility of Extraction Procedure for Alkaloids in Tobacco
Nicotine,
Nornicotine,
Total Alkaloid as Nicotine,
Nicotine,
h-ornicotine,
2.24 2.26
2.08 2.11
0.15 0.14
4.68 4.69
4.41 4.42
0.25 0.25
2.56 2.56
2.34 2.34
0.20 0.20
4.52 4.51
4.18 4.17
0.31 0.31
1.65 1.66
1.50 1.50
0.15 0.14
3.80 3.85
3.62 3.67
0.16 0.16
1.16 1.15
1.04 1.01
0.11 0.13
4.15 4.21
3.88 3.94
0.24 0.25
3.05 3.05
2.83 2.87
0.21 0.16
2.96 2.98
2.73 2.76
0.21 0.20
0.54 0.55
0.05 0.05
%
%
7Z
%
0.60 5.09 0.27 5.38 5.09 0.26 0.60 5.38 Standard deviation for total alkaloids = 0 . 0 1 7 Standard devjatjon for nicotine .= 0.021 Standard deviation for nornicotine = 0 . 0 1 2
%
%
with a fair degree of accuracy by direct titration of an aliquot of the hydrocarbon extract without removing the ammonia. Precision of Method. A detailed study of the precision and accuracy of the method was not made because only a limited amount of nornicotine was available for this investigation. Some measure of the precision was obtained by analyzing ten samples from each of two known aqueous nicotine solutions. One solution was prepared so that each aliquot contained 28.40 mg., the approximate amount of nicotine which would be found in the analysis of a low alkaloid tobacco. The average recovery for the ten analyses was 28.39 mg., with a standard deviation of 0.36. The second solution was prepared so that each aliquot contained 199.6 mg., the approximate amount of nicotine which would be found in the analysis of a high alkaloid tobacco. The average recovery for these ten analyses vias 199.30 mg. with a standard deviation of 0.50. I n addition, a flue-cured and burley tobacco sample were each analyzed ten times for total alkaloid content as nicotine by the described method. Results of these determinations are given in Table 111. Table IV indicates the precision obtsined on twelve duplicate analyses of burley tobacco samples for total alkaloid, nicotine, and nornicotine content. Comparison with Other Alkaloid Procedures. Table V compares the results for total alkaloids with those obtained by the AOAC silicotungstate method ( I ) . The Bowen-Barthel steam distillation apparatus ( 4 ) was used for alkaloid isolation in the A0.4C method. These data indicate that both methods are equally reliable. A comparison of results obtained by the Houston chromate graphic procedure (9) and the extraction procedure on a series of tobacco samples is shown in Table VI.
Table V. Total Alkaloids in Tobacco as Determined by Extraction and AOAC Silicotungstate lMethods
The potentiometric end point of the acetylated benzene-chloroform-acetic acid solutions was found to be -535 to -540 mv. with the methanol modified calomel electrode. This equivalence point is sufficiently reproducible to warrant the use of an automatic titrator, or of titrating to a definite millivoltage with standard titrators. Effect of Ammonia. The presence of ammonia in the hydrocarbon extract would lead to erroneous results in the nicotinenornicotine determination. It was believed that any ammonia absorbed during the extraction could be removed by aspirating the hydrocarbon solution for a short time prior to titration. Such aspiration would probably not remove any of the alkaloids. To test this and to ascertain the magnitude of the error due to the presence of ammonia, the following steps were taken. Known solutions of ammonium sulfate were made alkaline then extracted with benzene-chloroform solution. One aliquot of the extract was titrated directly; the second aliquot was transferred to a 125-ml. Erlenmeyer flask and a gentle stream of air was passed over the surface of the solution for 5 minutes, then titrated. In addition, known amounts of nicotine and nornicotine were treated in the same manner to ascertain whether aspiration would cause volatilization of either alkaloid. Approximately 1 to 2% of the added ammonia was extracted by the benzene-chloroform solution and this was effectively removed by the specified aspiration technique. None of the alkaloids was volatilized in a 5minute aspiration period. Ammonia will react quantitatively with acetic anhydride to yield acetamide, a nonbaeic compound in the nonaqueous media, but since acetic anhydride will also react with nornicotine, acetylation would not be an effective means of eliminating ammonia as an interferant in this particular analysis. Because such a small quantity of ammonia is absorbed by the extracting solution, total alkaloids as nicotine can be determined
Per Cent Total Alkaloids as Nicotine Extraction AOAC method method 6.03 5.99 4.71 4.74 3.12 3.15 3.60 3.59 n ti7 0.60 i.22 1.16 3.53 3.52 2.97 2.95
Table VI.
Comparison of Extraction Procedure with the Houston Chromatographic Procedure
Extraction Procedure, Nornicotine 0.16 0.25 0.37 0.45 0.17 0.21 1.34 1.44
Nicotine 1.68 2.85 4.30 5.54 2.96 2.85 0.43 0.39
% Total 1.84 3.10 4.67 5.99 3.13 3.06 1.77 1.83
Houston Procedure, % Nornicotine Total 0 27 1.88 0 38 3.16 0.46 4.62 0.39 6.04 5.05 0.36 3.11 2.75 2.71 3.10 0.39 0.38 1.63 1.28 0.35 1.38 1.73
Nicotine 1.61 2.78 4.16
DISCUSSION
The method has been used successfully in this laboratory for more than 2 years and has been found to offer advantages of simplicity and specificity over a number of other alkaloid procedures in use a t present. Some difficulty was experienced in obtaining reproducible results if the barium hydroxide was too finely ground. The reason for this is not readily apparent; therefore, it is recommended that the barium hydroxide be used as obtained from suppliers. Nicotine and nornicotine were found to be dibasic in all of the nonaqueous systems investigated. A perchlorate precipitate separated in each of the hydrocarbon solutions when titrated with
V O L U M E 27, NO. 10, O C T O B E R 1 9 5 5 perchloric acid, but did not interfere with the discernment of the equivalence point. The addition of acetonitrile to the titration solutions prevented precipitate formation; however, as commercial grade acetonitrile is slightly basic, it was necessary to apply a blank correction when it was used. I n analyzing tobacco samples the indicator end point was found to be sufficiently sharp not to warrant the continued use of acetonitrile. However, acetonitrile addition to the titration solutions would be advant:igeous in the assay of commercial nicotine solutions. It is believed that this procedure offers several advantages as a routine procedure for alkaloid analyses. All of the alkaloids are removed from the tobacco in a 15-minute extraction period, the hydrocarbon extract may be titrated directly, thus eliminating the acid extraction and indirect titration steps necessary in other titration procedures, no distillation is required as in the gravimetric and spectrophotometric procedures, ammonia may be eliminated as an interfering rnaterial in the analysis, and a means is provided for determining tertiary and secondary alkaloids in the same solution. ACKNOWLEDGMENT
The authors wish to express their appreciation to Abner Eisner, Eastern Regional Research Laboratories, for a supply of nornicotine picrate; to Forrest Houston, University of Kentucky, for furnishing some of the samples and analyses used in this study; to Frank B. Wilkinson, Tobacco Branch of the Agricultural Marketing Service, U. S. Department of Agriculture, for permission to use data obtained on tiamples submitted by his committee; to Joel P. Clingman and Herman H. hlusselwhite for the silicotungstate analyses; and to Jacquelyn B. Conrad for her asyistance in the precision studies.
1653 LITERATURE CITED (1) Assoc. Offic. Agr. Chemists, “Official 1Iethods of Analysis,” 7th ed., p. 69, 1950. (2) Avens, A. W., and Pearce, G . W., IND. ESG. CHEX,Axar.. ED., 11, 505 ( 1 9 3 9 ) . (3) Bertrand, G.. and Javillier, AI., B t d . soc. c h i m . F r u m e , 5 , 241 ( 1 9 0 9 ) . (4) Rowen, C. V.,and Barthel, W. F., ISD.EXG.CHEhf., ASAL. ED.,15, 596 (1943). (5) Ibid., p. 740. (6) Chapin, R. 11.,C . S.Dept. Agr., Bur. Animal Ind., Bull. 133 (1911).
W. W.,Bacon. C. W.,Bowling, J . D., and Brown, D . E., U. S.D e p t . -4gr., Tech. Bull. 414, 35 (1934). (8) Griffith, K. B., and Jeffrey, R. K.,~ ~ X A LCHEII., . 20, 307 (7) Garner,
(1948).
Houston, F. G., Ibid.,2 4 , 1831 (1952). (IO) Jeffrey, R. K.,J . Assoc. Oflc.Agr. Chemists, 34, 843 (1951). (11) Kissling, R., U. S. D e p t . Agr., Bur. Chem.. BiiZZ. 107, rev. 32 (9)
(1912); 2. anal. Chem., 2 1 , 64 (1882). (12)
Larson, P. S.,and Haag, H. R., IXD.ESG. CHEX.,.IN.AL. ED.,
(13) (14) (15)
Narkwood, L. N., J . Assoc. Ofic. Agr. Chemists, 2 2 , 427 (1939). Ibid., 2 6 , 283 (1943). Ogg, C. L., Willits, C. O., and Ricciuti, C. R., ANAL.CHEM.,22,
16, 86 (1944).
336 (1950). (16) Schloesing, >I., Compt. rend., 23, 1142 (1846). (17) Seaman, W..and Allen, E., ANAL.CHEM.,2 3 , 592 (1951). (18) Siggia, S.,Hanna, J. G., and Kervenski, I. R., Ibid., 22, 1295 (1950). (19) T o t h . J.. Chem.-Zto.. 25. 610 (1901). (205 Wagner, C. D., Brown, 11. H.;and’Peters. E. D . , J . Am. Chem. Soc., 69, 2609 (1947). (21) Willits, C . O., Swain, 11. L., Connelly, J. A., and Brice, B. A., ASAL. CHEM.,22, 430 ( 1 9 5 0 ) . (22) Wolff, W. A., Hawkins. 11. .1.,and Giles, W.E., J . B i d . Chem., 175, 825 (1948). RECEIVEDfor review January 3, 1955. .4ccepted May 21, 1955.
Vol umetric Determination of Zi iconium An Ethylenediamine Tetraacetate Method Involving Back-Titration with Bismuth JAMES S. FRITZ and MARLENE JOHNSON institute for A t o m i c Research and D e p a r t m e n t of Chemistry, /ora State C o l k g e , Amos, lowa Zirconium can be determined in acid solution by addition of excess ethylenediamine tetraacetate, followed by back titration with bismuth nitrate. Thiourea is used as the indicator. Verj few metals form complexes which are strong enough to interfere with the method. Most complexing anions, including sulfate, phosphate, oxalate, thiocyanate, and tartrate, do not interfere. Fluoride (except in low concentrations) interferes but can be removed easily by fuming w-ith perchloric or sulfuric acid. Slight modification of the procedure permits determination of zirconium in the presence of moderate amounts of thorium, titanium, niobium, or tantalum without separation.
F
OR many years macro quantities of zirconium have been determined by tedious, time-consuming gravimetric methods. Recently, however a number of titrimetric procedures have been proposed. Kolthoff and Johnson ( 4 ) reported an amperometric titration of zirconium, thorium, tin, and uranium with m-nitrophenylarsonic acid. White (8) developed an alkalimetric method for zirconium following the quantitative isolation of zirconium mandelate. Dhar and Das Gupta ( 1 ) determined zirconium by either colorinctric or volumetric means following precipitation with oxalohydroxamic acid. Fritz and Fulda (3)developed a
selective method in which zirconium is titrated u ith disodium ethylenediamine tetraacetate [disodium (ethylenedinitrilo) tetraacetate EDTA] using Eriochromecyanine indirator. Milner and Phennah ( 5 ) isolated the zirconium in zirconium-uranium alloys by means of mandelic acid, then added ewes5 E D T A and back-titrated the excess with ferric chloride using salicylic acid indicator. Very recently Olson and Elving ( 6 ) published a procedure for the amperometric titration of zirconium with cupferron. In the new method, excess E D T A is added to the sample, forming a stable complex with zirconium. The excess is then titrated 1%ith bisumuth nitrate using thiourea indicantor. The p H , indicator concentration, etc., are the same as described recently for the titration of bismuth (2). Few metal rations interfere because zirconium and bismuth form stronger complexes with E D T A than most metals, and in the back-titration bismuth does not react with the thiourea indicator until the metal-EDTA complexes which are much weaker than bismuth have been broken. Another important advantage is that zirconium can be determined in the preEence of such anions as phosphate, sulfate, thiocyanate, tartrate, and small amounts of fluoride. EFFECT O F AVIONS
Zirconium can be titrated in the presence of chloride, nitrate, perchlorate, tartrate, and thiocyanate without an\- difficulty
