USP 36 Official Monographs / Clorazepate 3067 Assay preparation; rU and rS are the peak responses obtained tain a solution having a known concentration of about from the Assay preparation and the Standard preparation, re- 75 µg per mL. Transfer 4.0 mL of this solution to a 50-mL spectively, M1 is the molecular weight of cloprostenol conical flask, add 4.0 mL of 0.7 M potassium carbonate, (424.92), and M2 is the molecular weight of cloprostenol 2.0 mL of Internal standard solution, and 15.0 mL of water. sodium (446.90). Insert a stopper, and mix. Test solutionÐTransfer an accurately weighed quantity of about 50 mg of Clorazepate Dipotassium to a 50-mL conical flask. Add 4.0 mL of 0.7 M potassium carbonate, and start . stirring the solution. Add 2 mL of Internal standard solution Clorazepate Dipotassium and 19.0 mL of water. Stop stirring about 5 minutes after the addition of the 0.7 M potassium carbonate solution. [NOTEÐPrepare fresh immediately before each injection.] Chromatographic system (see Chromatography 〈621〉)ÐThe liquid chromatograph is equipped with a 232-nm detector and a 3.9-mm × 30-cm column that contains packing L1. The flow rate is about 1.0 mL per minute. Chromatograph the Standard solution, and record the peak responses as di- rected for Procedure: the relative retention time for 2,6- C16H11ClK2N2O4 408.92 dimethylaniline is about 0.8 and 1.0 for nordazepam; the 1H-1,4-Benzodiazepine-3-carboxylic acid, 7-chloro-2,3- relative standard deviation of the peak area ratio of dihydro-2-oxo-5-phenyl-, potassium salt compound with nordazepam to 2,6-dimethylaniline for replicate injections is potassium hydroxide (1:1). not more than 2.0%. Potassium 7-chloro-2,3-dihydro-2-oxo-5-phenyl-1H-1,4- ProcedureÐSeparately inject equal volumes (about 20 µL) benzodiazepine-3-carboxylate compound with potassium of the Standard solution and the Test solution into the chro- hydroxide (1:1) [57109-90-7]. matograph, record the chromatograms, and measure the peak areas. Calculate the percentage of nordazepam in the » Clorazepate Dipotassium contains not less than portion of Clorazepate Dipotassium taken by the formula: 98.5 percent and not more than 101.5 percent of C16H11ClK2N2O4, calculated on the dried basis. 2500(C/W) (Ri / RS) Packaging and storageÐPreserve under nitrogen in in which C is the concentration, in mg per mL, of USP tight, light-resistant containers. Nordazepam RS in the Standard solution; W is the weight, in USP Reference standards 〈11〉Ð mg, of Clorazepate Dipotassium taken to prepare the Test USP 2-Amino-5-chlorobenzophenone RS solution; Ri is the peak area ratio of any impurity to 2,6- C13H10ClNO 231.68 dimethylaniline obtained from the Test solution; and RS is the USP Nordazepam RS peak area ratio of nordazepam to 2,6-dimethylaniline ob- 7-Chloro-1,3-dihydro-5-phenyl-2H-1,4-benzodiazepin- tained from the Standard solution: not more than 0.5% of 2-one. nordazepam is found and not more than 0.1% of any indi- C15H11ClN2O 270.72 vidual impurity is found. USP Clorazepate Dipotassium RS TEST 2Ð IdentificationÐ DiluentÐPrepare a mixture of 0.001 N sodium hydroxide A: Infrared Absorption 〈197M〉. and acetonitrile (1:1). B: Ultraviolet Absorption 〈197U〉Ð Mobile phaseÐPrepare a filtered and degassed mixture of Solution: 7 µg per mL. water, acetonitrile, and a 1 M solution of tetrabutylammo- nium hydroxide in methanol (110:90:1), adjust with phos- Medium: sodium hydroxide solution (1 in 2500). phoric acid to a pH of 7.7, and mix. Make adjustments if Loss on drying 〈731〉ÐDry it in vacuum at 60° for 1 hour: necessary (see System Suitability under Chromatography it loses not more than 0.5% of its weight. 〈621〉). Heavy metals, Method II 〈231〉: 0.002%. Standard solutionÐDissolve an accurately weighed quan- Related compoundsÐ tity of USP 2-Amino-5-chlorobenzophenone RS in Diluent, TEST 1Ð and dilute quantitatively, and stepwise if necessary, with Di- luent, to obtain a solution having a known concentration of Phosphate buffer solutionÐDissolve about 13.8 g of mon- about 0.0026 mg per mL. obasic sodium phosphate in 500 mL of water, adjust with 2.5 N sodium hydroxide to a pH of 8.0, and mix. Test solutionÐTransfer about 300 mg of Clorazepate Di- potassium, accurately weighed, to a glass test tube. Add Mobile phaseÐPrepare a filtered and degassed mixture of 10.0 mL of Diluent, and vigorously mix on a vortex mixer water, acetonitrile, and Phosphate buffer solution (5:4:1). for about 90 seconds. [NOTEÐPrepare fresh immediately Make adjustments if necessary (see System Suitability under before each injection.] Chromatography 〈621〉). Chromatographic system (see Chromatography 〈621〉)ÐThe Internal standard solutionÐDissolve about 5 mL of 2,6- liquid chromatograph is equipped with a 238-nm detector dimethylaniline in 50 mL of hexane, and carefully add drop- and a 3.9-mm × 30-cm column that contains packing L1. wise hydrochloric acid to precipitate the amine hydrochlo- The flow rate is about 2 mL per minute. Chromatograph the ride. Filter through a sintered-glass funnel, wash the solid Standard solution, and record the peak responses as directed precipitate with hexane, and allow the precipitate to dry. for Procedure: the relative standard deviation of the peak Transfer about 50 mg of the dried precipitate of 2,6- height for replicate injections is not more than 3.0%. dimethylaniline hydrochloride to a 100-mL volumetric flask, add 10.0 mL of Phosphate buffer solution and 40 mL of ProcedureÐSeparately inject equal volumes (about 20 µL) water, and dilute with acetonitrile to volume. of the Standard solution and the Test solution into the chro- matograph, record the chromatograms, and measure the Standard solutionÐDissolve an accurately weighed quan- peak responses. Calculate the percentage of each impurity tity of USP Nordazepam RS in acetonitrile, and dilute quan- titatively, and stepwise if necessary, with acetonitrile to ob- 3068 Clorazepate / Official Monographs USP 36 in the portion of Clorazepate Dipotassium taken by the Test solutionÐTransfer 1 Tablet to a suitable container, formula: add 200 mL of 0.01 M sodium hydroxide, and homogenize for not less than 3 minutes. Centrifuge a portion of this so- 1000(C/W)(ri / rS) lution for 15 minutes, and filter the supernatant, discarding the first 20 mL. Dilute an accurately measured portion of the in which C is the concentration, in mg per mL, of USP filtrate with 0.01 M sodium hydroxide to obtain a solution 2-Amino-5-chlorobenzophenone RS in the Standard solution; having a known concentration of about 7.6 µg per mL. W is the weight, in mg, of sample taken; ri is the peak ProcedureÐConcomitantly determine the absorbances of height of each impurity obtained from the Test solution; and the Standard solution and the Test solution in 1-cm cells at rS is the peak height of 2-amino-5-chlorobenzophenone ob- the wavelength of maximum absorbance at about 231 nm, tained from the Standard solution: not more than 0.1% of with a suitable spectrophotometer, using 0.01 M sodium 2-amino-5-chlorobenzophenone is found, not more than hydroxide as the blank. 0.1% of any other individual impurity is found, and not more than 1.0% of total impurities in Test 1 and Test 2 is Related compoundsÐ found. METHOD IÐ AssayÐTransfer about 150 mg of Clorazepate Dipotassium, Phosphate buffer solutionÐDissolve about 13.8 g of mon- accurately weighed, to a 250-mL beaker, add 100 mL of obasic sodium phosphate in 500 mL of water, adjust with glacial acetic acid, and stir until dissolved. Titrate with 0.1 N 1 N sodium hydroxide to a pH of 8.0, and mix. perchloric acid VS, determining the endpoint potentiometri- Mobile phaseÐPrepare a filtered and degassed mixture of cally, using a glass electrode and a calomel electrode con- water, acetonitrile, and Phosphate buffer solution (5:4:1). taining a 1 in 100 solution of lithium perchlorate in glacial Make adjustments if necessary (see System Suitability under acetic acid. Perform a blank determination (see Titrimetry Chromatography 〈621〉). 〈541〉), and make any necessary correction. Each mL of Standard solutionÐDissolve an accurately weighed quan- 0.1 N perchloric acid is equivalent to 13.63 mg of tity of USP Nordazepam RS in acetonitrile, and dilute quan- C16H11ClK2N2O4. titatively, and stepwise if necessary, with acetonitrile to ob- tain a solution having a known concentration of about 66 µg per mL. Transfer 4.0 mL of this solution to a 25-mL volumetric flask, add 5.0 mL of 0.7 M potassium carbonate . and 3.0 mL of acetonitrile, dilute with water to volume, mix, Clorazepate Dipotassium Tablets and filter. Test solutionÐWeigh and finely powder not fewer than » Clorazepate Dipotassium Tablets contain not 20 Tablets. Transfer an accurately weighed portion of the less than 90.0 percent and not more than powder, equivalent to about 15 mg of clorazepate dipotas- sium, to a suitable container. Add 5 mL of acetonitrile, 5 mL 110.0 percent of clorazepate dipotassium of 0.7 M potassium carbonate, and 15 mL of water, stir for (C16H11ClK2N2O4). 10 minutes, and filter. [NOTEÐPrepare fresh before each in- jection, and use within 3 minutes.] Packaging and storageÐPreserve in tight, light-resistant containers. Chromatographic systemÐThe liquid chromatograph is equipped with a 232-nm detector and a 3.9-mm × 30-cm USP Reference standards 〈11〉Ð column that contains packing L1. The flow rate is about USP 2-Amino-5-chlorobenzophenone RS 1.5 mL per minute. Chromatograph the Standard solution, C13H10ClNO 231.68 and record the peak responses as directed for Procedure: the USP Nordazepam RS relative standard deviation for replicate injections is not 7-Chloro-1,3-dihydro-5-phenyl-2H-1,4-benzodiazepin- more than 2.0%.