Sulfanilamide, Sulfapyridine, Sulfathiazole, Sulfaguanidine, and Sulfadiazine Assay, Differentiation, and Identification J. A. CALAMARI, ROBERT HUBATA, AND P. B. ROTH Laboratory, New York Medical Depot, Brooklyn, N. Y.
Differentiation
sulfapyridine, sulfathiazole, sulfahave been studied by the authors, and methods for assay, differentiation, and identification have been established. Sulfanilamide has been adopted as a primary standard for the standardization of the nitrite solution used in the assay of the sulfa drugs, because of the ease with which it may be purified and its stability in air. Rapid differentiation of the sulfa drugs from each other is accomplished by means of the solubility of the original compounds and the solubility and color of some of the easily prepared derivatives. The identity of the sulfa drugs is established by determining the melting points of both the original drugs and the 2-amino hydrolysis products in the case of sulfapyridine, sulfathiazole, and sulfadiazine. The melting points of sulfanilamide and sulfaguanidine and the presence of ammonia in their hydrolysis products identify them. The authors have selected the diazotization reaction for the assay of the sulfa drugs in preference to an element analysis such as that depending on the estimation of sulfur (1), because the former is based on the presence of a characteristic group—the aryl amino group—and because of the greater rapidity of the assay. Using the easily purified sulfanilamide as a primary standard for the standardization of the sodium nitrite volumetric solution, the possibility of error is less than in assay methods employing volumetric solutions standardized by more indirect methods such as the sulfanilamide assay of the U. S. Pharmacopoeia (4). Furthermore, the assay method is applicable to all five sulfa drugs either as pure compounds or in tablets, and since the standardization reaction and procedure are identical with those of the assay method, the possibility of an end-point error is minimized. The information necessary for the differentiation and identification of the sulfa drugs is given in Table I. The methods employed in establishing these characteristics are described below, in following paragraphs numbered to correspond with those in the table. Table I shows that sulfanilamide can be differentiated from the other sulfa drugs by its solubility in water; sulfaguanidine by its insolubility in sodium hydroxide solution; sulfathiazole by the insolubility of its diazonium chloride at 0° to 4° C.; and sulfadiazine by the insoluble compound formed with stannous chloride.
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SULFANILAMIDE, guanidine, and sulfadiazine
Table I. Meltine Point.
0
0
C.
2. 3. 4.
Temperature of preheated bath, Solubility in 5% HC1 Solubility in 5% NaOH Solubility in water, mg. per 100
5. 6.
Basic hydrolysis product, melting point Diazonium chloride, color of solution and
7. 8. 9.
SnCb-HCl Solubility in acetone Solubility in ether
1.
solubility
(3).
Standardization.
Dissolve 0.52 gram of pure sulfanilamide in 7 cc. of hydrochloric acid, A. C. S., and 35 cc. of water. Add 30 grams of ice and titrate with 0.1 M sodium· nitrite volumetric solution, using starch iodide paper or paste as Each cubic centimeter of 0.1 M sodium an outside indicator. nitrite is equivalent to 0.01722 gram of sulfanilamide. (see reagents)
Identification
cc.
(S)
170°
Soluble Soluble Insoluble, 49.5 at 37° C.
Soluble Soluble Insoluble, 94 at 37° C.
Soluble Insoluble Insoluble, 190 at 37° C.
Yellow Soluble
56°
90° Yellow Insoluble
NHs Colorless Soluble
126° Pale yellow Soluble
No ppt. Soluble Insoluble
No ppt. Soluble Insoluble
No ppt. Soluble
White ppt. Soluble
191-192° (decomposed) 170°
150°
No ppt.
Insoluble
Sulfa Drugs
Sulfapyridine
Soluble Soluble Soluble. Very soluble in hot water. 800 at 25° C. NHs Colorless Soluble Soluble
of
Sulfadiazine 251-252° (decomposed) 225° Soluble Soluble Insoluble, 12.3 at·. 37° C.
Sulfanilamide 165.5-166.0°
C.
and Identification
Follow the U. S. P. XI melting point method (5), but preheating the bath to the temperature indicated in Table I, continuing heating at the rate of 0.5° per minute, and stirring vigorously with a propeller type of motor-driven stirrer. Sulfaguanidine is rendered anhydrous by drying at 110° C. prior to the melting point determination. 2. To 0.1 gram of the drug add 10 cc. of an approximately 5 per cent solution of hydrochloric acid, A. C. S., at 25° C. Stir to effect solution. 3. To 0.1 gram of the drug add a 5 per cent solution of sodium hydroxide, reagent grade, at 25° C. and stir. 4. To 0.1 gram of the drug add 20 cc. of distilled water at 25° C. and stir vigorously. The numerical values offered have been obtained from the literature (3). 5. Reflux 1 to 2 grams of the drug with 50 cc. of hydrochloric acid, A. C. S., for 1 to 2 hours. Cool and make strongly alkaline with a 25 per cent solution of sodium hydroxide. Cool and extract with ether. Collect the ether extractions, filter, and evaporate. Determine the melting point by the U. S. P. XI (6) method, employing vigorous stirring as in paragraph 1. 6. Place 0.50 gram of the drug in a test tube and dissolve in 7 cc. of hydrochloric acid, A. C. Si, and 35 cc. of distilled water. Cool in an ice water bath. Add 20 cc. of 0.1 M sodium nitrite solution slowly with constant stirring and cooling. Cool the contents of the tube to 0° to 4° C. 7. Dissolve 0.1 gram of the drug in 5 cc. of a 5 per cent solution of hydrochloric acid, A. C. S., and add 1 cc. of stannous chloride-hydrochloric acid reagent. Cool to 20° C. and allow to stand for 10 minutes. 8. Add 20 cc. of acetone, A. C. S., to 0.1 gram of the drug and agitate. 9. Add 20 cc. of ether, U. S. P., to 0.1 gram of the drug and agitate. Reagents. Sodium nitrite solution. Dissolve 7.0 grams of sodium nitrite, A. C. S., to make 1 liter of 0.1 N or 0.1 M solution. Standardize as directed below. Stannous chloride-hydrochloric acid reagent. Dissolve 50 grams of stannous chloride, reagent grade, in 50 cc. of hydrochloric acid, A. C. S. Sulfanilamide (for primary standard). Recrystallize once from acetone and twice from water. Starch iodide paper, U. S. P. XI (7), and starch iodide paste 1.
534
Sulfathiazole 201-202°
Sulfaguanidine 190.5-191.5°
(decomposed) 180°
0-4°
at
Insoluble
Insoluble
ANALYTICAL
July 15, 1942
EDITION
535
Assay
Discussion
The assay procedure is identical with that employed in the standardization. If the approximate quantity of the sulfa drug is not known, run an exploratory titration as in standarization, using the outside indicator after each 1-cc. addition of sodium nitrite. On the final titration, weigh a sample calculated to yield an approximate 30-cc. titration, dissolve, cool the sample as before, and add the sodium nitrite to within about 1 or 2 cc. of the value obtained in the exploratory titration, then in intervals of 0.1 cc., and as the end point is approached in 0.05 and finally 0.025 cc., testing with the outside indicator after each addition. Stir vigorously during and after each addition of sodium nitrite. Tablets of the sulfa drugs contain 5 or 7.7 grains of the drugs and therefore smaller quantities than 30 cc. of sodium nitrite will be consumed if the tablets are assayed individually. The accuracy is proportional to the quantity of sodium nitrite solution consumed.
Sodium nitrite solutions standardized against purified sulfanilamide and standardized against sodium oxalate by the U. S. P. sodium nitrite assay method (5) have agreed within 0.1 per cent. Sulfathiazole, sulfaguanidine, and sulfadiazine purified by repeated crystallization from acetone have assayed 100.0 =±=0.1 per cent and recrystallized sulfapyridine 0.1 per cent, using the diazotization method. Tablets 99.9 have been assayed directly by the method described and the sulfa drugs have been separated from the tablet excipients by extraction and recrystallization for determination of the other characteristics given in Table I.
If the quantity of sample indicated below is used, of about 30 cc. will be obtained: Quantity of Sulfanilamide Sulfapyridine Sulfathiazole Sulfaguanidine, anhydrous Sulfadiazine
Sample Gram 0.52
0.75 0.75 0.64 0.75
a
titration
Each Cc. of 0.1 N
NaNOi Equivalent to: .Gram 0.01722 0.02492 0.02553 0.02142 0.02501
=±=
Literature Cited (1) Am. Medical Assoc., “New and Non-Official Remedies”, 1940
ed., p. 492. (2) Ibid., 1941 ed., pp. 518-19. (3) Roblin, R. O., Williams, J. H., Winneck, P. S., English, J. P., J. Am. Chem. Soc., 62, 2002 (1940). (4) United States Pharmacopoeia XI, 2nd Supplement, p. 101, (5) United States Pharmacopoeia XI, p. 344. (6) Ibid., p. 455. (7) Ibid., p. 561.
Rapid Detection of Gold by the Electrographic Method J. A. CALAMARI, ROBERT HUBATA, and P. B. ROTH Laboratory, New York Medical Depot, Brooklyn, N. Y.
in alloys and in plating may be detected rapidly by tests (8-7) and by electrographic methods (1), the latter being the more rapid. The authors have discovered a rapid test for gold in alloys and plating which may be completed in about one second and is apparently specific for gold. When gold is made the anode in a neutral or slightly acid solution of nitrate ion, using an e. m. f. of 6 to 9 volts and a current of about 0.5 to 1.0 ampere, it enters solution as Au+++ and reacts with water to form auric hydroxide which deposits on the anode. Hydrogen peroxide added to the solution reduces the auric hydroxide to purple aurous hydroxide and colloidal gold.
GOLD spot
Apparatus and Test Solutions. The equipment and procedure are identical with those described for the electrographic test (2). The test solution is prepared by dissolving 30 grams of sodium nitrate, reagent grade, in sufficient hydrogen peroxide, U. S. P., to make 100 ml., and filtering. Method. Ashless filter paper is folded to form a wad three to four layers thick and then dipped into the sodium nitrate-hydrogen peroxide solution. The moistened filter paper is placed on the test metal anode and a graphite cathode about 7.5 cm. (3 inches) long and 0.6 cm. (0.25 inch) in diameter is firmly applied to the free side of the moistened section of the filter paper for about one second. An e. m. f. of about 6 to 9 volts is employed and may be obtained by using 4 to 6 No. 6 dry cell batteries connected in series. The current is about 0.5 to 1 ampere. An e. m. f. of 4.5 to 6 volts, a current flow of 0.1 to 0.5 ampere, and about a 5-second contact are recommended for plating.
If gold is present, a purple stain appears on the paper adjacent to the test metal anode. The intensity of the color increases with the percentage of gold present in the test metal when current density and time remain constant. The stain also increases in intensity upon standing. Very thin or porous plating yields a faint pui pie to purple stain; heavy plating, a purple stain.
A strong positive test for gold has been obtained with a series of dental alloys ranging from 25 per cent, to pure gold, and with 14- and 18-karat jewelry golds. Copper and silver
in the gold alloys tested did not interfere. Numerous other metals and alloys have been tested by this method and negative results have been obtained in each case: nickel-silver, platinum-ruthenium, platinum-iridium, solders, brasses, white metals, 15 per cent silicon steel, bronzes, copper-nickel alloys, carbon steels, platinum, palladium, nickel, copper, manganese, molybdenum, tantalum, tungsten, mercury, cadmium, aluminum, tin, zinc, vanadium, silver, and lead. Chromium yields a blue spot which fades rapidly. Vanadium yields a red spot and silver a black spot. It is believed that the test is applicable to alloys containing less than 25 per cent of gold, because the purple stain obtained with the alloys containing 25 to 30 per cent of gold is intense. Heavy gold plating can be detected readily. Thin or porous gold plate on sterling silver yields a spot, faint purple to purple in color, often interspersed with dark areas, due to silver. The characteristic gold stain, however, has been clearly discernible in all tests performed. Very thin gold plate on copper or brass cannot be detected by this method.
Literature Cited (1) Arnold, E„ Chem. Listy, 27, 73-8 (1933). (2) Calamar!, J. A., Ind. Eng. Chem., Anal. Ed., 13, 19-20 (1941). (3) Costeanu, R. N., Z. anal. Chem., 104, 351-5 (1936). (4) Duval, C., and Fauconnier, P., Mikrochim. Acta, 3, 30-2 (1938). (5) Feigl, F., Krumholz, P., and Rajmann, E., Mikrochemie, 3,
165-73 (1931).
(6) Holzer, A., Ibid., 8, 275 (1930). (7) Tananaev, N. A., and Dolgov, K. A., J. Russ. Phys.-Chem. Soc.,, 61, 1377-84 (1929).
