Analyst, August, 1974, Vol. 99, pp. 487-490 487 Colorimetric Determination of Antazoline in Some Pharmaceutical Preparations with Sodium Nitrite BY M. M. AMER, M. S. TAWAKKOL (Department of Analytical Chemistry, Faculty of Pharmacy, Cairo University, Cairo, Egypt) AND s. A. ISMAIEL (Research Department, Soci&te' Misr pour l'lndustrie Pharmaceutique, 92 El Mataria Street, Post El Zeitoun, Cairo, Egypt) A colorimetric method for the determination of antazoline in pharma- ceutical preparations with sodium nitrite has been developed. The method involves treatment of a cooled and acidified dilute aqueous solution of the sample with sodium nitrite. The yellow colour produced is stabilised by the addition of propan-2-01 or ethanol and the absorbance then measured at 410 nm. Naphazoline, tolazoline, clemizole, diphenhydramine, chlorpheniramine, ephedrine, cetrimide, benzalkonium chloride and zinc salts, even if present in amounts ten times greater than that of antazoline, do not interfere.ANTAZOLINE can be determined in some pharmaceutical preparations by using the acidimetric methods of the B.P.,l the B.P.C.,2 the non-aqueous titration methods of the U.S.P.,3 Selles and Rodriguez4 and Rink and F3emhofer5 or by using the EiZ& value mentioned by Clarke,6 but none of these methods can be used if antazoline occurs in combination with ephedrine, diphenhydramine or chlorpheniramine. Phenylephrine, if present, interfered in the determination by the last four of the above methods, while naphazoline interfered in the determination by the methods of Slack and Mader,' Horioka and Ishiokas and K ~ m - T a t t , ~ giving high results.In this paper, a sensitive colorimetric method for the determination of antazoline in pharmaceutical preparations is proposed in which use is made of the qualitative colour reaction reported by Auterhoff.lO The proposed method was found to be simple, quick and devoid of the drawbacks of the methods mentioned above. EXPERIMENTAL REAGENTS- Antazoline hydrochloride standard stock solution-Dissolve 100 mg of antazoline hydro- chloride in sufficient water to give a final volume of 100 ml. Antazoline methanesulphonate standard stock solution-Dissolve an amount of an tazoline methanesulphonate (previously determined according to the B.P.C. 1968) in sufficient water to give a final concentration of 100 mg of antazoline methanesulphonate per 100 ml.Sodium nitrite solution-Dissolve 2 g of sodium nitrite (Merck) in sufficient water to give a final volume of 100 ml. Hydrochloric acid, approximately 2.5 N. Hydrochloric acid, concentrated. Ethanol, 95 per cent. CALIBRATION GRAPH- Dilute the appropriate volume of the antazoline salt stock solution with water to give solutions of final concentrations ranging between 0.4 and 2-4mg of the antazoline salt per 100 ml. Transfer 5 ml of the diluted antazoline salt solution (containing 20 to 120 pg of the corresponding antazoline salt) into a glass-stoppered test-tube, cool it in ice, add 2 ml of 2.5 N hydrochloric acid, mix and then add 1 ml of sodium nitrite. Mix the solution and, after @ SAC and the authors.488 [Analyst, Vol.99 2 minutes, add 2 ml of ethanol, mix again, then remove the test-tube from the ice and, after 3 minutes, measure, at 410 nm, the absorbance of the yellow colour produced against a blank carried out simultaneously. AMER et aZ. : COLORIMETRIC DETERMINATION OF ANTAZOLINE PREPARATION OF SAMPLE SOLUTIONS- Tablets-Transfer an amount of the powder containing about 50 mg of antazoline hydro- chloride into a 100-ml calibrated flask, add about 80 ml of water and shake the flask until the powder has dissolved (about 30 minutes). Dilute this solution to volume with water, filter it through a dry Whatman No. 1 filter-paper and dilute 2 ml of it to 100 ml with water. Drops, injectable solutions and syru$s-Dilute an appropriate volume of the sample with water so as to give a final concentration of about 1 mg of the antazoline salt per 100 ml.Creams-Transfer an amount of the cream containing about 100 mg of antazoline hydro- chloride into a beaker and extract it successively with 50, 20 and 20 ml of 0.5 N hydrochloric acid by heating the mixture each time on a boiling water bath, while stirring, until the cream has melted, allowing it to stand for about 5 minutes, then cooling it in ice, and decanting the aqueous extract into a separating funnel. Extract the combined aqueous extracts with about 10 ml of chloroform, make the aqueous layer up to 200 ml with water, then dilute 2 ml of the latter to 100 ml with water. Lotions-Transfer 5 ml of the well shaken sample into a 100-ml calibrated flask, add 5 ml of concentrated hydrochloric acid, mix and, when the effervescence ceases, dilute the mixture to volume with water.Filter it through a dry Whatman No. 1 filter-paper and reject the first turbid 10 ml of the filtrate, then dilute a volume containing about 1 mg of antazoline hydrochloride to 100 ml with water. DEVELOPMENT OF THE COLOUR- Transfer 5 ml of the prepared sample solution into a glass-stoppered test-tube and complete the procedure as directed under Calibration graph, starting with “cool it in ice, add 2 ml of 2.5 N hydrochloric acid. . .” Calculate the concentration of the antazoline salt from the calibration graph, RESULTS AND DISCUSSION The results obtained by applying the proposed method to the determination of antazoline hydrochloride and antazoline methanesulphonate in some pharmaceutical preparations are given in Table I.TABLE I COMPARISON OF RESULTS BY THE PROPOSED METHOD AND SLACK AND MADER’S METHOD Antazoline salt I i Pharmaceutical Indicated Added Recovery, per cent.? preparations* on to f A > label/ labelled by proposed by Slack and Mader’s material / mg method method Sensol tablets 100 - 99.6 f 1.06 103-04 f 1-34 Antistine ampoules 50 - 100.4 f 1.14 101.5 f 1.89 Antistine privine solution 500 - 98.42 f 2.32: 112 f 2.521 Fenozal drops5 500 100-92 f 0.61 101-92 f 0.31 Calazol lotion 1000 - 95.6 f 1 not applicable Calazol cream 2000 - 105 f 2.37 not applicable mg - 50 100.1 f 0.65 25 100 f 0.81 - 250 98.93 f 1.35 - 500 97.4 & 0.82 1000 103.83 f 1.82 - * Sensol tablets, fenozal drops, calazol lotion and calazol cream were supplied by Socidtd Misr pour Antistine ampoules and antistine privine solution were supplied ? Mean of six experiments.$ Calculated as antazoline hydrochloride ; for conversion into antazoline sulphate results were multiplied 4 Specially prepared authentic sample containing the correct amount of ingredients. 1’Industrie Pharmaceutique, Cairo, Egypt. by Ciba Laboratories, Basle, Switzerland. by 1.041.August, 19741 IN SOME PHARMACEUTICAL PREPARATIONS WITH SODIUM NITRITE 489 The qualitative colour reaction of antazoline and sodium nitrite reported by Auterhoff lo was studied and used in order to develop a quantitative method for the determination of antazoline in pharmaceutical preparations. The absorption spectrum of the colour obtained showed a maximum at 410 nm (Fig.1) and the maximum intensity of the colour was obtained by using 20mg of sodium nitrite (Fig. 2) and hydrochloric acid of about 2.5 N concentration (Fig. 3) (both sulphuric and hydrochloric acids at about 2.5 N concentration gave the same colour intensity). The colour reached its maximum intensity after about 10 minutes, remained stable for about 5 minutes and then began to fade (Fig. 4). Wavelength/nrn Fig, 1. Absorption spectrum of the colour produced 0.4 ~ 0.3 2 0.2 Fi 0 Is m 0.1 I I I I I I I J 0 5 10 15 20 25 30 35 40 Sod i u rn nit r it e/mg Fig. 2, Effect of the amount of sodium nitrite on the intensity of the colour The use of ethanol (or propan-2-01) was found to improve the stability of the colour (Fig. 4), the intensity of which obeyed Beer's law for amounts of antazoline hydrochloride between 20 and 120 pg and of antazoline methanesulphonate between 20 and 140 pg.The colour was found to be unstable at room temperature or on heating the solution but cooling the latter in ice or to a temperature below 10 "C improved the stability of the colour (Fig. 4). As shown in Table I, the proposed method gave more accurate results than those obtained by the method of Slack and Mader, especially for samples that contain naphazoline. Diphenhydramine, chlorpheniramine, naphazoline, ephedrine, codeine, benzalkonium chloride, phenylephrine, tolazoline, clemizole, phenol, camphor, esters of p-hydroxybenzoic acid, sodium cyclamate, saccharin sodium, zinc oxide and calamine in amounts ten times greater than that of antazoline did not interfere.0 1 2 3 4 Hydrochloric acid/N Fig. 3. Effect of concentration of hydrochloric acid on the intensity of the coIour490 AMER, TAWAKKOL AND ISMAIEL The reaction is probably due to the aniline moiety of the antazoline molecule, because naphazoline and tolazoline, which contain the same imidazoline ring but no aniline moiety, did not give a colour, whereas phentolamine and diethylaniline, which contain an aniline moiety, gave a yellow colour with a maximum absorption at 360 nm. 0.4 I 1 0 5 15 25 35 Time elapsed/minutes Fig. 4. Improved stability of the colour a t temperatures below 10 “C (solutions cooled in an ice-cold water-bath for both stable and un- stable colours) : A, ethanol - water; and B, water The explanation that the colour results from the formation of a nitroso derivative of antazoline similar to that which occurs in the nitrosation of a dialkylaniline such as diethyl- aniline with nitrous acid, as stated by Finarl11l2 and Mann and Saunders,l39l4 cannot be supported as the maximum absorption of the yellow colour produced with phentolamine and diethylaniline differed greatly from that with antazoline.1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. REFERENCES “British Pharmacopoeia 1968,” The Pharmaceutical Press, London, 1968, p. 56. “British Pharmaceutical Codex 1968,” The Pharmaceutical Press, London, 1968, p. 46. “United States Pharmacopeia,” Fifteenth Revision, 1955, Mack Publishing Company, Easton, Pa., Selles, E., and Rodriguez, A. M., Galenica Acta, 1956, 9, 33; Chem. Abstr., 1957, 15, 8375 h. Rink, M., and Riemhofer, M., Mitt. dt. pharm. Ges., 1961, 31, 197. Clarke, E. G. C., assisted by Berle, J., “Isolation and Identification of Drugs in Pharmaceuticals, Slack, S. C., and Mader, W. J., J . Amer. Pharm. Ass., Sci. Edn., 1957, 46, 742. Horioka, M., and Ishioka, H., J . Pharm. SOC. Japan, 1961, 18, 76. Kum-Tatt, L., J . Pharm. Pharmac., 1960, 12, 666. Auterhoff, H., Arch. Pharm., Bed., 1950, 283, 244. Finar, I. L., “Organic Chemistry, The Fundamental Principles,” Volume I, Fifth Edition, The p. 62. Body Fluids and Post Mortem Material,” The Pharmaceutical Press, London, 1969, p. 198. English Language Book Society and Longman Group Limited, London, 1969, p. 621. -, oP. cit., p. 697. Mann, F. G., and Saunders, B. C., “Practical Organic Chemistry,” Longmans, Green and Co. Ltd., Fourth Edition, London, 1960, p. 202. -__ , , op. cit., p. 376. Received December 17th, 1973 Accepted February 25t12, 1974