Table 11. Oxygen in Aluminum Electrolytically Refined Aluminum
Energy Irradiaafter tion etchings, time, MeV min 11.8 11.8
20 20
11.5
35
11.1 10.7
20 20
10.0 8.2 8.2
20 15
12.0
20 15 60
15
Chemical separation Without Elimination of Na on hydrated SbzOj Steam distillation of HzSiF, Without Elimination of Na on hydrated SbzOs Without Without Without
Oxygen content, Pg/g 0.23 i- 0.02 0 , 1 9 =t0.02 0.23 f 0.02
0 . 2 0 i- 0.02 0.23 & 0 . 0 2 0.20 i 0.02 0.21 i. 0 . 0 2 0.23 i. 0.02
Zone Refined Aluminum 11.6 11.4
Without Without Without
50.04 20.04
ing to sodium-24. The sensitivity limit for the detection of fluorine-18 has been calculated considering that the activity due to this element is lower than twice the statistic fluctuation of the activity measurements of sodium-24. Twice the highest value of this fluctuation is considered to be the activity limit of fluorine-18. The sensitivity of the analysis of oxygen in aluminum samples obtained from zone melting can undoubtedly be increased by the removal of sodium since the statistics fluctuation will then be decreased. The sodium-24 can be removed by retention on hydrated antimony pentoxide. We have also performed some determinations of oxygen in aluminum from a single electrolysis (nominal purity 99.7 %) with and without chemical separation of fluorine. The results obtained vary between 1 and 20 pg/gram; these values depend on the importance of the chemical etching. The variation in the apparent contents of oxygen could be due t o the irregular chemical attack on samples of aluminum with a high content of impurities.
20.05
CONCLUSION Sodium can be separated by its retention o n hydrated antimony pentoxide (19) after dissolving the sample in hydrochloric acid (6N). Fluorine, partially retained in the absence of aluminum (20), is completely eluted in the presence of this element. We have confirmed, by steam distillation of hydrofluoric acid at 200 “C (ZI), that the period of 110 minutes really corresponds to fluorine-18 (Figure 3). RESULTS
The oxygen content was calculated using the so-called “equivalent thickness” method (22) and the activation curve determined by Markowitz. The results obtained are presented in Table 11. The only detectible half-life in the case of the purest aluminum samples (zone refined aluminum) is 15 hours correspond(19) F. Girardiand E. Sabbioni,J. Radioanal. Chem., 1, 169(1968). (20) B. Vialatte, J. N. Barrandon, S . Alexandrov, I. N. Bourrelly, C. Cleyrergue, N. Deschamps, and H. Jaffrezic, Radiochern. Radioanal. Lett., 5 (l), 59 (1970). (21) B. Vialatte, Bull. SOC.Chim. France, 1971, 347. (22) Ch. Engelmann, C.R. Acad. Sci., 258, 4279 (1964).
Because of the high sensitivity of activation analysis by SHe and the possibility of eliminating the effect of surface oxygen, we have been able to show that the oxygen content of double electrolysis aluminum (99.999 %) is some tenth micrograms/gram of Al. Such low contents of oxygen cannot be determined by other more classical methods of analysis. We could not detect any I8F in samples of higher purity (melting zone aluminum). After irradiation of 30 minutes in a beam of aHe with 2 PA intensity, we detected a limit of 0.05 pg of O/gram of AI. It would be possible to decrease this limit by irradiation with tritons Of higher energy (some l o MeV) since at 3 MeV only the cross-section of the reaction ‘SO(T,n)l8F is 0.8 barn (23). Unfortunately tritons Of this energy are not yet available for this kind Of
for review
26, lg71* Accepted October 26,
1971. (23) J. N. Barrandon and Ph. Albert, Rev. Phys. Appl., 3, 111 (1968).
New Method for Determination of 21-Hydroxy Corticosteroids Sandor Gorog and Gabor Szepesi Chemical Works G. Richter, Budapest 10, Hungary
THESTANDARD
METHODS of the corticosteroid analysis cannot differentiate between free 21-hydroxy compounds and their esters and hence these methods are not suitable for the determination of the 21-hydroxy contamination in 21-acyloxy corticosteroids. Tetrazolium reagents which are most frequently used for the determination of corticosteroids react only with the free 21-hydroxy derivative, but their 21-acyloxy derivatives hydrolyze under the strongly alkaline conditions employed in this method and hence they are also measured. The rate of color development of the two kinds of steroids is of course different. This is the basis of the differential
kinetic method of Guttman (1) which is suitable for the determination of free 21-hydroxy contamination over 1 %. Neither the original form of the Porter-Silber method ( 2 ) nor its modification by Lewbart and Mattox (3) is a suitable solution to the above problem. In this paper a new method is described for the specific determination of free corticosteroids based on spectrophotometry of their quinoxaline derivatives. (1) D. E. Guttman, J . Pharm. Sci., 55, 919 (1966). (2) C. C. Porter and R. H. Silber, J. Bid. Chem., 185, 201 (1950). ( 3 ) M. L. Lewbart and V. R. Mattox, ANAL.CHEM., 33, 559 (1961). ANALYTICAL CHEMISTRY, VOL. 44, NO. 6, M A Y 1972
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Table I.
Spectral Data of Prednisolone after Treatment with Aromatic Diamine Reagents I
Maximum, nm 318
Molar Medium absorptivity methanol 7470 1N HCl (see “Experimental”) 331 10200 i 1.1%a 4,5-Dimethyl methanol 331 7500 o-phenylene 1N HC1 351 12700 =t0.9%. diamine a Relative standard deviation calculated from six parallel runs.
Reagent o-Phenylene diamine
Table 11. Determination of Hydrocortisone in Hydrocortisone 21-Acetate
Taken, 0.20
z
Hydrocortisone Found, 0.22
0.41 0.83 1.24
z
0.41 0.84
1 . 2 1 + 0.017a 1.65 1.65 2.06 2.05 2.46 2.44 a Standard deviation calculated from six parallel runs.
EXPERIMENTAL Reagents. Cupric acetate reagent, 0.7 %. Dissolve 0.7 gram of cupric acetate in 10 ml of water and dilute the solution with methanol to 100 ml. 0-Phenylene diamine reagent, 0.1 %. Dissolve 0.1 gram of o-phenylene diamine in 5 ml of INhydrochloric acid and dilute t o 100 ml with water. 4,5-Djmethyl o-phenylene diamine was prepared according to Szhntay and Rohhly ( 4 ) . The reagent is prepared as described above. Procedure. Dissolve a n accurately weighed quantity of the sample, which contains about 1 mg of free 21-hydroxy corticosteroid in 25 ml of methanol. (In the determination of free 21-hydroxy corticosteroids in their 21-acyloxy esters, use 50 to 100 mg of the ester.) Add 5.0 ml of cupric acetate reagent. Allow the mixture to stand at room temperature for 1 hour, then add 1 ml of 1 N hydrochloric acid, 5.0 ml of o-phenylene diamine or 4,5-dimethyl o-phenylene diamine reagent and allow to stand for 1 hour again. Add 20 ml of 5N hydrochloric acid and dilute to volume with water. Determine the absorbance of this solution against a similarly treated reagent blank at 331 nm in the case of the o-phenylene diamine reagent or at 351 nm in the case of the dimethyl derivative. Calculate the 21-hydroxy corticosteroid content on the basis of the absorbance of a similarly treated standard solution. RESULTS AND DISCUSSION Determination of Optimum Conditions for the Reaction. Lewbart and Mattox (3) showed that corticosteroids could easily be oxidized by cupric acetate to the corresponding glyoxal derivative. This reaction serves as the first step of our method. The glyoxal derivative thus obtained is condensed with o-phenylene diamine o r its 4,Sdimethyl derivative to form the stable, spectrophotometrically active quinoxaline derivative. (4) Cs. Szdntay and J. RohBly, Period. Politech., Chem. Etzg., 8, 9
(1964). 1080
ANALYTICAL CHEMISTRY, VOL. 44, NO. 6, MAY 1972
As 21-acyloxy corticosteroids d o not react with cupric acetate, this procedure is specific to free corticosteroids. We have found that under the conditions described under “Experimental,” both the oxidation and the condensation reaction are nearly quantitative. This can be proved by comparing the data of Table I with those described in the literature. Leanza et al. (5) and Lewbart and Mattox (6) found molar absorptivities of 7950 and 8150, respectively, at 319 nm for crystalline quinoxaline derivatives in neutral methanolic media. The oxidation by cupric acetate is carried out in neutral methanolic medium, similar to the procedure of Lewbart and Mattox (3). We have found that the excess of cupric acetate does not disturb either the formation of quinoxaline derivative o r the spectrophotometric measurement, and hence both reactions can be carried out successively without any separation. Mild acidic medium proved to be suitable for the condensation reaction. Both o-phenylene diamine and its 4,5-dimethyl derivative have been tried in this step of the procedure. They have been found equivalent from the viewpoint of reactivity, chromophore stability, and linearity. The spectrophotometric measurement is carried out in strongly acidic medium (1N hydrochloric acid) where the quinoxaline derivative exists in fully protonized state. As can be seen in Table I, the acidification of the solution results in a bathochromic shift and increased sensitivity. A further advantage of the strongly acidic medium is that the excess of the diamine reagents is also protonized and this results in low absorbance of the reagent blank at the analytical wavelength. From among the maxima of the spectrum of quinoxaline de. ivatives, the long wavelength one has been chosen for the measurements. The more intense short wavelength peak (5,6) cannot be used because of the strong absorbance of the excess reagent. I n 1 N hydrochloric acid, the chromophor is stable for at least 24 hours. Beer’s law has been found valid within the 0.01-1 .O absorbance range. Although o-phenylene diamine has the advantage of being commercially available, we have used in practice the dimethyl derivative because of its more favorable spectral characteristics. As seen in Table I, the introduction of the methyl groups in the molecule results in a bathochromic and hyperchromic effect. The former effect is advantageous in some special cases (assay of formulated corticosteroids) while the latter increases the sensitivity of the method to be nearly equal to the triphenyl tetrazolium method. (5) W. J. Leanza, J. P. Conbere, E. F. Rogers, and K. Pfister, J . Amer. Chem. SOC.,76,1691 (1954). (6) M. L. Lewbart and V. R. Mattox, J . Urg. Chem., 28, 2001 ( 1963).
Scope and Limitations. The method seems to be suitable for the determination of any kinds of steroids with a dihydroxy acetone side chain a t position 17. The position and strength of the absorption band is independent of the structure and substituents of the steroid skeleton. The absorption maxima of the dimethyl quinoxaline derivatives for prednisolone, prednisone, hydrocortisone, cortisone, triamcinolone etc. were found a t 351 nm and the molar absorptivities range between 12050 and 12700. The above mentioned corticosteroids can be determined as contaminants in their 21 -acylated derivatives (acetates, trimethylacetates, p-toluene sulfonates, methane sulfonates, etc.) if their quantity is more than 0.2 %. The data of Table I1 are characteristic of these investigations. The application of the method t o 21-amino corticosteroids and pharmaceutical formulations will be published later.
17-Deoxy corticosteroids cannot be investigated by the proposed method because in this case the 20-keto-21-aldehyde side chain which is formed in the oxidation step, is instantaneously transformed t o the stable enol-aldehyde form (17/20/-ene-20-01-2l-a1) and the latter does not react with ophenylene diamine. It must be noted that triamcinolone acetonide cannot be determined also. The reason for this is supposed t o be the steric hindrance caused by the bulky acetonide group. Precision of the Method. The relative standard deviations in Tables I and I1 are characteristic of the precision of the method.
RECEIVED for review August 9, 1971. Accepted November 3, 1971. This paper is the 20th in a series on Analysis of Steroids.
Separation of Cadmium(l1) from Zinc(l1) and Other Metal Ions on a Cadmium Selective Exchanger: Titanium Selenite Mohsin Qureshi, R a j e n d r a Kumar, a n d H. S. R a t h o r e Department of Chemistry, Aligarh Muslim Unhersity, Aligarh, U. P., India
THEMOST OBVIOUS advances in ion exchange in the last few years are in the area of inorganic ion exchangers ( I ) . Numerous new materials have been reported and older ion exchange materials have been studied in greater detail. Most applications of ion exchange to chemical analysis are chromatographic and ion exchange chromatography can be used to separate metal ions. Of the various metal ions, cadmium is especially important since most of the methods for determining this element depend o n the complete removal of almost all other elements ( 2 ) . Several ion exchange methods for the separation of cadmium from zinc o n resins in hydrochloric acid (3), aliphatic alcohols (4,dimethyl sulfoxide-aqueous hydrochloric acid ( 5 ) and in hydrobromic acid (6) media have been reported. It has been mentioned (7) that these complicated and costly solvents can be replaced by aqueous solutions containing inorganic solutes by using synthetic inorganic ion exchangers instead of organic resins. The selectivity of a n ion exchanger depends largely on its composition. Numerous ion exchangers were thoroughly studied but none of them was effective for this separation. We have therefore investigated a new ion exchanger, titanium selenite. The unique feature of this material is its high selectivity for cadmium. In this report we summarize a simple and rapid method for the separation of cadmium from zinc and from numerous metal ions by the use of a titanium selenite column. (1) H. F. Walton, ANAL.CHEM., 42, 86R (1970). (2) I. M. Kolthoff and P. J. Elving, “Treatise on Analytical Chemistry.” Vol. 3. Interscience, New York, N.Y., 1961, p 183. (3) H. G. Meyer, Z . Arid. Cliem., 24, 394 (1969). (4) F. W. E. Strelow, C. R. Vanzyl, and C. J. C. Bothma, A m / . Cliim. Acra, 45, 81 (1969). ( 5 ) I. Brize, L. W. Marple, and H. Diehl, Talama, 15, 1441 (1968). (6) J. Korkisch and E. Klakl, ibid., 16, 377 (1969). (7) G. Alberti, “Chromatography Review,” Vol. 8, Elsevier, New York, N.Y., 1966, p 246.
EXPERIMENTAL
Apparatus. All instrumentation was the same as that used in our earlier papers (8-10). Reagents. Sodium selenite (Reidel), selenious acid (BDH), and 15% wt/v titanic chloride (BDH) were used. All other chemicals were of analytical grade. Method of Preparation. Samples 2, 3, and 6 were prepared by mixing the 0.10M titanic chloride and sodium selenite solutions in the volume ratio of 1:2 at p H -1.20, 9.90, and 6.70, respectively. For sample 7, the mixing ratio was 1 : 1 ; the rest of the conditions were similar t o sample 2. Sample 1 was prepared by mixing 0.025M solutions of titanic chloride and sodium selenite in the volume ratio 1 :2 at p H -2. Sample 4 was prepared by refluxing a-titanic acid in 0.20M selenious acid for 36 hr. I n order t o obtain sample 5, equal volumes of 0.2M solutions of titanic chloride and selenious acid were mixed and the product obtained was refluxed for 36 hr. Ammonia solution (d = 0.88) was used fo1 raising the p H wherever necessary. The precipitates were washed, filtered, dried, converted t o hydrogen form, sieved t o 60-120 mesh, washed again with demineralized water, and finally dried at 50 “C. The exchanger was now ready for use. The Kd values, the saturation ion-exchange capacity for K-, the pH-titration curves, X-ray, and IR studies were made as reported earlier (8, 9). For conductometric and high frequency titrations, 2 ml of 0.10M sodium selenite solutions were taken in the cell, diluted t o 100 ml with water, and titrated with 0.10M titanic chloride solution. Column Operation. For separation studies, titanium selenite (2.0 grams) .was filled in a glass column (0.68 i d . ) with glass wool support, the column was washed with de(8) M. Qureshi, H. s. Rathore, and R. Kumar, J . Cliem. Sac. ( A ) , 1970, 1986. (9) M. Qureshi, H. S. Rathore, and R. Kumar, J. Cliromatogr., 1971, 269. (10) M. Qureshi and J. P. Gupta, J . Cliem. Soc. ( A ) , 1969, 1755. ANALYTICAL CHEMISTRY, VOL. 44,
NO. 6, M A Y 1972
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