EFAVIRENZ Final text for addition to The International Pharmacopoeia

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1 Document QAS/05.145/FIAL March 07 EFAVIREZ Final text for addition to The International Pharmacopoeia This monograph was adopted at the Fortieth W Expert ommittee on Specifications for Pharmaceutical Preparations in ctober 2005 for addition to the 4 th edition of the International Pharmacopoeia. EFAVIREZUM EFAVIREZ Relative molecular mass hemical name. (4S)-6-chloro-4-(cyclopropylethynyl)-1,4-dihydro-4-(trifluoromethyl)-2-3,1- benzoxazin-2-one. AS Description. White to slightly pink powder. Solubility. practically insoluble in water, freely soluble in methanol. ategory. Antiretroviral (on-nucleoside Reverse Transcriptase Inhibitor). Storage. Efavirenz should be kept in a well-closed container, protected from light.

2 Working document QAS/05.145/FIAL07 page 2 Additional information. Efavirenz may exhibit polymorphism. Requirements Efavirenz contains not less than 97.0% and not more than 103.0% of calculated with reference to the dried substance. Manufacture. The production method is validated to ensure that the substance is the 4S enantiomer. Identity test Either tests A,B and D or tests and D may be applied A. arry out test A.1. or, where UV detection is not available, test A.2. A.1. arry out the test as described under Thin-layer chromatography, using silica gel R6 as the coating substance and a mixture of 90 volumes of dichloromethane R, 10 volumes of methanol R and 3 volumes of glacial acetic acid R as the mobile phase. Apply separately to the plate 5 µl of each of 2 solutions in methanol R containing (A) 5 mg of the test substance per ml and (B) 5 mg of efavirenz RS per ml. After removing the plate from the chromatographic chamber, allow it to dry exhaustively in air or in a current of cool air. Examine the chromatogram in ultraviolet light (254 nm). The principal spot obtained with solution A corresponds in position, appearance, and intensity with that obtained with solution B. A.2. arry out the test as described under Thin-layer chromatography, using silica gel R5 as the coating substance and a mixture of 90 volumes of dichloromethane R, 10 volumes of methanol R and 3 volumes of glacial acetic acid R as the mobile phase. Apply separately to the plate 5 µl of each of 2 solutions in methanol R containing (A) 5 mg of the test substance per ml and (B) 5 mg of efavirenz RS per ml. After removing the plate from the chromatographic chamber, allow it to dry exhaustively in air or in a current of cool air. Dip the plate in basic potassium permanganate (~1 g/l) TS. Examine the chromatogram in daylight. The principal spot obtained with solution A corresponds in position, appearance, and intensity with that obtained with solution B. B. The absorption spectrum of the final solution prepared for the Assay, when observed between 210 nm and 300 nm, exhibits one maximum at about 247 nm; the specific absorbance (A 1% 1cm) is between 449 to arry out the examination as described under 1.7 Spectrophotometry in the infrared region. The infrared absorption spectrum is concordant with the spectrum obtained from efavirenz RS or with the reference spectrum of efavirenz. If the spectra thus obtained are not concordant, repeat the test using the residues obtained by separately dissolving the test substance and efavirenz RS in a small

3 Working document QAS/05.145/Final07 page 3 amount of methanol R and evaporating to dryness. The infrared absorption spectrum is concordant with the spectrum obtained from efavirenz RS. D. Determine the specific optical rotation using a 3 mg/ml solution in methanol and calculate with 20 reference to the dried substance; [ α ] = 89 to 100. D Sulfated ash. ot more than 2.0 mg/g. Loss on drying. Dry for 4 hours at 105 ; it loses not more than 5 mg/g. Related substances Prepare fresh solutions and perform the tests without delay. arry out the test as described under igh-performance liquid chromatography, using a stainless steel column (15cm x 4.6mm, 3.5 µm), packed with a cyanopropyldimethylsilane monolayer (e.g. Zorbax SB-). The mobile phases for gradient elution consist of a mixture of Mobile phase A and Mobile phase B, using the following conditions: Mobile phase A: 90 vol of a 0.05% solution of trifluoroacetic acid R and 10 vol of methanolr. Mobile phase B: 10 vol of a 0.05% solution of trifluoroacetic acid R and 90 vol of methanolr. Time (min) Mobile phase A (%) Mobile phase B (%) Prepare the following solutions in a mixture of 50 % acetonitrile and 50 % water (PL quality) (dissolution solvent). For solution (1) dissolve 25 mg of the test substance in the dissolution solvent and dilute to 25.0 ml with the same solvent. For solution (2) dilute 1.0 ml of solution (1) to 50.0 ml with the dissolution solvent and dilute 5.0 ml of the resulting solution to ml with the same solvent. For solution (3) dissolve 5 mg of efavirenz RS in 5 ml of a solution prepared as follows: dissolve 1 mg of efavirenz impurity B RS in the dissolution solvent and dilute to 10 ml with the same solvent, dilute 1 ml of the resulting solution to 25 ml with the same solvent. perate with a flow rate of 1.5 ml per minute. As a detector use an ultraviolet spectrophotometer set at a wavelength of about 250 nm. Inject separately 35 µl each of solutions (1), (2) and (3) and of the dissolution solvent in the

4 Working document QAS/05.145/FIAL07 page 4 chromatographic system. Examine the blank chromatogram for any extraneous peaks and disregard the corresponding peaks observed in the chromatogram obtained with solution (1). In the chromatogram obtained with solution (3), the impurity B peak is eluted at a relative retention of about 0.9 with reference to efavirenz (retention time about 20 minutes).the test is not valid unless the resolution between impurity B and efavirenz is at least 3. In the chromatogram obtained with solution (1) the area of any peak corresponding to impurity B is not greater than four times the area of the principal peak in the chromatogram obtained with solution (2) (0.4%), the area of any other peak, apart from the principal peak, is not greater than twice the area of the principal peak in the chromatogram obtained with solution (2) (0.2%) and the area of not more than three such peaks is greater than the area of the principal peak in the chromatogram obtained with solution (2) (0.1 %). The sum of the areas of all peaks, other than the principal peak, is not greater than 8 times the area of the principal peak in the chromatogram obtained with solution (2) (0.8%). Disregard any peak with an area less than 0.5 times the area of the principal peak in the chromatogram obtained with solution (2) (0.05%). Assay Dissolve about 25 mg, accurately weighed, in methanol to produce 50.0 ml; dilute 1.0 ml of this solution to 50.0 ml with the same solvent. Measure the absorbance of a 1-cm layer of the diluted solution at the maximum at about 247 nm. alculate the amount of 14 9 I 2 using an 1% absorptivity value of 47.3 (specific absorbance: A 473). 1 cm =

5 Working document QAS/05.145/Final07 page 5 Impurities Me A. -[4-chloro-2-[(1S)-3-cyclopropyl-1-hydroxy-1-(trifluoromethyl)prop-2-ynyl]phenyl]-4- methoxybenzamide B. (4S)-6-chloro-4-[(1E)-2-cyclopropylethenyl]-4-(trifluoromethyl)-1,4-dihydro-2-3,1- benzoxazin-2-one, 3. (4S)-6-chloro-4-(pent-1-ynyl)-4-(trifluoromethyl)-1,4-dihydro-2-3,1-benzoxazin-2-one,

6 Working document QAS/05.145/FIAL07 page 6 3 D. (4S)-6-chloro-4-(2-cyclopropylethynyl)-1-(4-methoxybenzyl)-4-(trifluoromethyl)-1,4- dihydro-2-3,1-benzoxazin-2-one, R E. R = : (2S)-2-(2-amino-5-chlorophenyl)-4-cyclopropyl-1,1,1-trifluorobut-3-yn-2-ol, F. 6-chloro-2-cyclopropyl-4-(trifluoromethyl)quinoline,

7 Working document QAS/05.145/Final07 page 7 3 G. (4S)-6-chloro-4-[2-(2-methylcyclopropyl)ethynyl]-4-(trifluoromethyl)-1,4-dihydro-2-3,1- benzoxazin-2-one, (enantiomer) [see under Manufacture] 2 3. methyl [4-chloro-2-[(1S)-3-cyclopropyl-1-hydroxy-1-(trifluoromethyl)prop-2- ynyl]phenyl]carbamate, Me I. (2S)-2-[5-chloro-2-[(4-methoxybenzyl)amino]phenyl]-4-cyclopropyl-1,1,1-trifluorobut-3-yn- 2-ol, Me J. (4S)-6-chloro-4-(2-cyclopropylethynyl)-2-(4-methoxyphenyl)-4-(trifluoromethyl)-1,4- dihydro-2-3,1-benzoxazine.

8 Working document QAS/05.145/FIAL07 page 8 Reagents Basic potassium permanganate (~1g/l) TS A solution of potassium permanganate R containing about 1 g of KMn 4 in 1 litre of sodium hydroxide (~40g/l)TS. Sodium hydroxide (~40g/l) TS A solution of sodium hydroxide R containing about 40 g/l of a (approximately 1 mol/l). Triflouroacetic acid 2 2 A commercially available reagent of suitable grade. ***