from a phase-lock loop. Consequently, the only electronic device not constructed from integrated circuits could be a strip-chart recorder.
ACKNOWLEDGMENT The authors thank Dorys Olivares for her assistance in the preliminary stages of this work. The authors are also indebted to the following people for their aid in the design and construction of parts of the apparatus: M. Williams, K. Bastin, R. Ensman, and M. Lockto. We are also grateful for the use of the nebulizer-burner system supplied by VarianTechtron.
LITERATURE CITED (1) (2) (3) (4)
E. Berman, Appl. Spectrosc., 29, 1 (1975). W. Snelleman, Spectrochirn. Acta, Part B, 23,403 (1968). V. Svoboda. Anal. Chern., 40, 1384 (1968). R. C. Elser and J. D. Winefordner. Anal. Chem., 44, 698 (1972)
(5) G. J. Nitis. V. Svoboda, and J. D. Winefordner, Spectrochirn. Acta, Part 8, 27, 345 (1972). (6) W. Snelleman. T. C. Rains, K. W. Yee, H. D. Cook, and 0. Menis, Anal. Chern., 42, 394 (1970). (7) D. G. Mitchell, J. M. Rankin. and E. W. Bailey. Spectrosc. Lett.. 5, 37 (1972). (8) I. S.Maines. D. G. Mitchell, J. M. Rankin, and E. W. Bailey, Spectrosc. Len., 5, 251 (1972). (9) G. M. Hieftje and R. J. Sydor, Appl. Spectrosc., 26, 624 (1972). (IO) R. Mavrodineanu and H. Boiteux, "Flame Spectroscopy," J. Wiley and Sons, New York, 1965, p 371. (11) G. M. Hieftje, Anal. Chem., 44 (6), 81A (1972). (12) G. M. Hieftje and R. i, Bystroff. Spectrochim. Acta. Part B, 30, 187 (1975). (13) G. Horlick and K. R. Betty, Anal. Chem., 47, 363 (1975).
RECEIVEDfor review November 3, 1975. Accepted November 21, 1975. The authors acknowledge support of this study through National Science Foundation Grant MPS75-21695. Reference to a company or product name does not imply approval or recommendation of the product to the exclusion of others which might be suitable.
Spectrophotometric Determination of Niobium(V) as a Mixed Ligand Complex with Gallic Acid and 1,2Diaminocyclohexanetetraacetic Acid M. Subbarao and K. Srinivasulu' School of Studies in Chemistry, Vikrarn University, Ujjain (M.P.),lndia
Niobium(V) was determined as a mixed ligand complex with gallic acid and 1,2-diaminocyclohexanetetraacetlc acid in tartrate medium by a spectrophotometric method at 470 nm around pH 2.5, 40 minutes after preparation of the solution and at 25 'C. The system conforms to Beer's law for an optimum range of 0.38-8.92 pg/ml of niobium with relative standard deviation of 2 to 5 % . Sensitivity ( A = 0.001 and 1-cm path length) is 0.015 pg/mi of niobium. The behavior of many ions was studied for their interference in determination of niobium and, in a few synthetic cation mixtures, niobium was estimated.
Few mixed ligand complexes of niobium and tantalum were studied for their stoichiometry, stability constants ( I , 2 ) and analytical determinations (3, 4 ) including extraction procedures (5). Gallic acid complex of niobium in the presence of 1,2-diaminocyclohexanetetraacetic acid (DCTA) was studied in tartrate medium. Optimum conditions for its analytical determination in pure form and in the presence of foreign ions including a few synthetic mixtures were established and described below.
EXPERIMENTAL A Bausch and Lomb Spectronic 20 spectrophotometer using K in. matched cells and a Systronics (India) pH meter (Type 322-1) were used. Niobium(V) solution M) (Specpure Nb205, Johnson Mathey & Co., London) in 0.75% tartaric acid, after fusing the oxide in 1g of potassium bisulfate, was prepared. A solution of 1,2-diaminocyclohexanetetraaceticacid (DCTA) (K & K Labs, Inc. USA) M) was also prepared with a few drops of sodium hydroxide t o give a clear solution. Freshly prepared aqueous solutions of gallic acid (10-1M) (recrystallized pure sample, Riedal, Germany); 0.75% of tartaric acid
solution, and titanium and tantalum in 0.75% tartaric acid solutions were used. Ten ml of gallic acid, 2.7 ml of DCTA, and varying amounts of niobium in tartrate medium were mixed; the p H adjusted a t 2.5 f 0.1, and made up to 25 ml with distilled water of the same pH. The final tartrate concentration was maintained within 1.0 to 1.3 ml of tartaric acid solution including the niobium (in tartrate medium) concentration taken. Absorbance measurements were taken a t 470 nm, 40 minutes after preparation of the solution a t 25 "C.
RESULTS AND DISCUSSION Effect of Reagent Concentration. Niobium(V) in tartrate medium forms a yellow colored complex with gallic acid with an absorption maximum a t 390 nm and a colorless solution with DCTA alone. But when mixed with both the ligands in tartrate medium (but not in oxalate medium), it forms an orange-red colored complex with absorption maximum a t 470 nm. When gallic acid in the range of 10 to 15 ml, DCTA in the range of 2.5 to 3.0 ml, are mixed with 1 ml of niobium solution a t p H 2.5 f 0.1 in a total volume of 25 ml a t room temperature, and when over-all tartaric acid concentration is maintained a t 1 to 1.3 ml including niobium solution taken, absorbance values a t 470 nm remained the same if measured after 40 minutes of preparation of solutions. This complex persists only in the 2.1 to 2.7 pH range and when the temperature does not exceed 30 'C but is stable for several hours. Beyond these ranges, the color intensity varies. Beer's Law, Range, Sensitivity, and Precision. The optimum niobium concentration range for its estimation and measurement a t 470 nm using %-in. matched cells was found to be 0.38 pg/ml to 8.92 pg/ml of niobium, and conformity to Beer's law was obtained. Sensitivity of the estimation is 0.015 pg/ml of niobium for A = 0.001 absorbance unit and 1.0-cm path length. Relative standard deviation of calculated absortivities of 28 samples in the optimum concentration range is 4.6%. ANALYTICAL CHEMISTRY, VOL. 48, NO. 3, MARCH 1976
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Study of Foreign Ions. About 120 times larger amounts of uranium(VI), 60 times of thorium(IV), 140 times of sodium and potassium, 35 times of rare earths(”, 20 times tungsten(VI), yttrium(III), ammonium; 18 times of zinc(II), 1 2 times of strontium(II), 10 times of copper(I1) and zirconium(IV), 5 times of lead(II), 3 times of antimony(V), tantalum(\’), iron(III), equal amounts of titanium(1V) and 50 times of nitrate, 40 times of chloride and phosphate, 140 times of sulfate did not show significant interference in the estimation of 37.6 pm of niobium in 25 ml by the recommended procedure. However, in the presence of large amounts (greater than 10 times) of zirconium and (i), in presence of 4 to 5 times of tantalum(V), iron(III), aluminum(III), fluoride ion; (ii) in the presence of comparable amounts of nickel(II), tin(I1) and (IV), beryllium(I1); and (iii) in the presence of small amounts of molybdenum( VI), this method gives errors in the range of 10%. The interference of 9 times of iron(II1) could be suppressed completely using ascorbic acid, but for molybdenum(V1) using citric acid as a masking agents, niobium could be estimated with 3% of error by this method. In the following synthetic mixtures in 25 ml of solution, niobium(V) was estimated successfully from (1) niobium (40 pg) with zirconium(1V) (91 pg) and uranium(V1) 3808 pg); (2) niobium(V) (37.16 pg) with zirconium(1V) (4.56 pg)
and uranium (VI) (190 pg); (3) Nb205 (44.59 pg); TazO5 (8.05 pg); Ti02 (1.62 pg); Si02 (0.22 pg); SnOz (1.48 pg); Fez03 (14.77 pg); CaO (0.94 pg); MgO (0.03 pg); A1203 (0.79 pg); Tho2 (1.21 pg); Crz03 (0.81 pg); YzO3 (13.93 p g ) ; MnO (1.60 p g ) ; PbO (2.04 pg); UOz (6.03 pg); NazO (0.29 pg); K20 (0.21 pg) (relative standard deviation of 7 samples is 2.6%); (4) Niobium(V) (92.9 pg) tantalum(V) (90.8 pg); titanium(1V) (11.0 pg); tin(I1) (50 wg); antimony(II1) (50 wg), and tungsten(V1) (800 pg). Niobium estimation by this method in the presence of other metals as given above will be useful for the analysis of fuel elements or other types of alloys where uranium(V1) concentration is high.
LITERATURE CITED (1) I. D. Ali-zade and 0 . A. Gamid-zade, Zh, Anal. Khim., 29,735-9 (1974). (2)A. K. Babko and V. V. Lukachine, Ukr. Khem. Zh., 27,682-7 (1961). (3)V . P. Madhava Menon, N. Mahadevan, K. Srinivasulu, and Ch. Venkateswarulu, J. Sci. hd. Res. (Hardwar, India), 218, 20-23 (1962). (4)V . Patrovsky, Collect. Trav. Chim. Tchec., 23 1774 (1958). (5) S.V. Elinson, L. T. Pobedina, and A. T. Rezova, Zavod. Lab., 37, 391-4
(1971).
RECEIVEDfor review August 21,1975. Accepted December 1, 1975. Financial assistance from the Council of Scientific and Industrial Research, India, to one of the authors (MSR) as an award of Junior Research Fellowship is gratefully acknowledged.
Colorimetric Assay for Aromatic Amines Esther Rinde and Walter Troll” New York University Medical Center, Department of Environmental Medicine, 550 First Avenue, New York, N. Y. 100 16
Aromatic amines can be detected in the nanomole range with the reagent Fluram, with which they form stable yellow derivatives. Fluram In glacial acetic acid reacts only with aromatic amlnes. The reaction is complete in 10 minutes and can be performed on thin-layer (TLC) chromatograms making possible the specific measurement of aromatic amines. The yellow product can be quantitatively eluted from the TLC plates. Fluram is colorless, and the blanks are zero.
The reagent Fluorescamine (Fluram) was introduced by Udenfriend (1) for the quantitation of aliphatic amines in a sensitive fluorescent assay. Aromatic amines, like the aliphatics, form fluorescent products with Fluram which are unstable, but they also form stable yellow derivatives. A number of aromatic amines have been found t o be carcinogens ( 2 ) ;hence, it is important to have sensitive and specific assays for their detection in the environment and in body fluids. Using selective extraction procedures ( 3 ) , the excretion of aromatic amine in the urine of exposed individuals can be measured, and is a good criterion for determining whether exposure has occured. The stability of the product formed when Fluram reacts with aromatic amines makes it possible to perform the assay on TLC from which it can be quantitatively eluted. Moreover by the use of TLC in conjunction with the assay, it is also possible to differentiate one aromatic amine from another ( 4 ) . One of the colorimetric assays we had previously used was a modi542
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fication of the Satake method ( 5 ) using the reagent trinitrobenzene sulfonic acid (TNBS) which reacts with aromatic amines a t pH 5. With TNBS, it is necessary to extract the product formed into organic solvent; otherwise the yellow color of the reagent interferes. Fluram has the advantage of being colorless (blanks are zero), eliminating the need for extraction, and has a sensitivity in the nanomole range.
EXPERIMENTAL Apparatus and Reagents. All solvents were Reagent Grade. Aniline, purchased from Eastman Organic Chemicals was redistilled 2 X . Purified samples of the other amines tested were supplied by Allied Chemical Company: benzidine, 2-naphthylamine, dichlorobenzidine, 0-tolidine. Fluram was purchased from Fisher Scientific (Catalog No. 43023). Monoacetyl benzidine was synthesized from benzidine (6). All of the aromatic amines were dissolved in acetone to give a solution (dichlorobenzidine, because of its limited solubility had to be prepared by first dissolving it in glacial acetic acid (glac. HAC), then diluting with acetone to this concentration). Fluram was used as a 1 mg/ml solution in glacial acetic acid which is stable for weeks a t room temperature. For the aliquoting of microliter quantities, the “Drummond Dialamatic Microdispenser” (Drummond Sci. Glass P6295) was used. Thin-layer chromatography (TLC) plates were purchased from Scientific Glass (Silica Gel 0.25 mm thick. Art 5762/0001). The solvent system for development of the plates was chloroform (90) glac. HAC (5) methanol (5). Spraying was accomplished with the “Lab Reagent Sprayer” (Analtech Catalogue No. A-100). Procedure. Fluram Reaction in Solution. Aliquots of 2 to 50 ~1 were transferred to tubes and the acetone was evaporated with a stream of nitrogen. Fifty gl of the Fluram solution was then added
