Chlordiazepoxide Hydrochloride and Clidinium Bromide Capsules
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This article is compiled based on the United States Pharmacopeia (USP) – 2025 Edition
Issued and maintained by the United States Pharmacopeial Convention (USP)
1 DEFINITION
Chlordiazepoxide Hydrochloride and Clidinium Bromide Capsules contain NLT 90.0% and NMT 110.0% of the labeled amounts of chlordiazepoxide hydrochloride (C16H14ClN3O · HCl) and clidinium bromide (C22H26BrNO3).
2 IDENTIFICATION
A. The retention times of the major peaks of the Sample solution correspond to those of the Standard solution, as obtained in the Assay.
3 ASSAY
3.1 PROCEDURE
[NOTE—Use low-actinic glassware.]
Buffer: Dissolve 1.92 g of sodium 1-pentanesulfonate in 900 mL of water in a 1-L volumetric flask. Adjust with 1 N sulfuric acid to a pH of 3.8 ± 0.1. Dilute with water to volume.
Mobile phase: Methanol, tetrahy drofuran, and Buffer (6:24:70) Diluent: Methanol and water (1:1)
Standard solution: 0.1 mg/mL of USP Chlordiazepoxide Hydrochloride RS and 0.05 mg/mL of USP Clidinium Bromide RS in Diluent Sample solution: Weigh the contents of NLT 20 Capsules, and calculate the average weight per Capsule. Mix the combined contents of the
Capsules, and transfer an amount equivalent to about 5 mg of chlordiazepoxide hydrochloride (C H ClN O · HCl ) to a 50-mL volumetric flask. Add about 25 mL of Diluent, sonicate for 5 min, and shake by mechanical means for 10 min. Dilute with Diluent to volume, and filter, discarding the first 20 mL of the filtrate.
3.1.1 Chromatographic system
(See Chromatography 〈621〉, System Suitability.) Mode: LC
Detector: UV 212 nm
Column: 8-mm × 10-cm; packing L1 Flow rate: 3 mL/min
Injection size: 20 µL
3.1.2 System suitability
Sample: Standard solution
[NOTE—The relative retention times for clidinium bromide and chlordiazepoxide hydrochloride are about 0.5 and 1.0, respectively.] Suitability requirements
Resolution: NLT 5.0 between the clidinium bromide and chlordiazepoxide hydrochloride peaks
Relative standard deviation: NMT 2.0% Analysis
Samples: Standard solution and Sample solution
Calculate the percentage of the labeled amount of chlordiazepoxide hydrochloride (C H ClN O · HCl) in the portion of Capsules taken:
Result = (ru/rs) × (Cs/Cu) × 100
ru = peak response of chlordiazepoxide hydrochloride from the Sample solution
rs = peak response of chlordiazepoxide hydrochloride from the Standard solution
Cs = concentration of USP Chlordiazepoxide Hy drochloride RS in the Standard solution (mg/mL)
Cu = nominal concentration of chlordiazepoxide hydrochloride in the Sample solution (mg/mL)
Calculate the percentage of the labeled amount of clidinium bromide (C H BrNO ) in the portion of Capsules taken:
Result = (rU/rS) × (CS/CU) × 100
= peak response of clidinium bromide from the Sample solution
= peak response of clidinium bromide from the Standard solution
= concentration of USP Clidinium Bromide RS in the Standard solution (mg/mL)
= nominal concentration of clidinium bromide in the Sample solution (mg/mL)
Acceptance criteria: 90.0%–110.0%
4 PERFORMANCE TESTS
Change to read:
DISSOLUTION ▲▲ (RB 5-NOV-2021) 〈711〉
Test 1: Use Dissolution 〈711〉, Procedure, Apparatus 1 and Apparatus 2, Immediate-release dosage forms, Procedure for a pooled sample for immediate-release dosage forms.▲ (RB 5-Nov-2021)
Medium: Water; 900 mL Apparatus 1: 100 rpm
Time: 30 min
Buffer: Dissolve 1.92 g of sodium 1-pentanesulfonate in 900 mL of water in a 1-L volumetric flask. Adjust with dilute sulfuric acid to a pH of
3.8 ± 0.1. Dilute with water to volume.
Mobile phase: Methanol, tetrahydrofuran, and Buffer (6:18:75)
Standard solution: Prepare a solution having known concentrations of USP Chlordiazepoxide Hy drochloride RS and USP Clidinium Bromide RS in Medium.
Sample solution: Pass a portion of the solution under test through a suitable filter. Combine equal volumes of the filtered solutions and use
the pooled sample for the analysis. Dilute with Medium to a concentration that is similar to that of the Standard solution, if necessary.
Chromatographic system
(See Chromatography 〈621〉, System Suitability.) Mode: LC
Detector: UV 212 nm
Column: 4-mm × 25-cm; packing L1 Flow rate: 2 mL/min
Injection size: 100 µL System suitability
Sample: Standard solution
[NOTE—The relative retention times for clidinium bromide and chlordiazepoxide hydrochloride are about 0.6 and 1.0, respectively.] Suitability requirements
Resolution: NLT 5.0 between the clidinium bromide and chlordiazepoxide hydrochloride peaks Relative standard deviation: NMT 2.0%
Analysis
Samples: Standard solution and Sample solution
Calculate the average percentage of chlordiazepoxide hydrochloride (C H ClN O · HCl) or clidinium bromide (C H BrNO ) dissolved: Result = (r /r ) × (C /L) × V × 100
= peak response of chlordiazepoxide hydrochloride or clidinium bromide from the Sample solution
= peak response of chlordiazepoxide hydrochloride or clidinium bromide from the Standard solution
= concentration of USP Chlordiazepoxide Hydrochloride RS or USP Clidinium Bromide RS in the Standard solution (mg/mL)
L = chlordiazepoxide hydrochloride or clidinium bromide label claim (mg)
V = volume of Medium (mL), 900
Tolerances: NLT 75% (Q ) each of the labeled amounts of chlordiazepoxide hydrochloride (C16H14ClN3O · HCl) and clidinium bromide (C22H26BrNO3) are dissolved.
Test 2: If the product complies with this test, the labeling indicates that it meets USP Dissolution Test 2.
Protect the Standard stock solution, Standard solution, and Sample solution from light. Medium: 0.1 N hydrochloric acid; 500 mL
Apparatus 1: 100 rpm
Time: 30 min
Buffer: Transfer 1.36 g of monobasic potassium phosphate to 1000 mL of water. Add 1.0 mL of triethylamine and adjust with 25% phosphoric acid to a pH of 3.6.
Mobile phase: Acetonitrile, methanol, and Buffer (25:10:65)
Standard stock solution: 0.32 mg/mL of USP Chlordiazepoxide Hydrochloride RS and 0.16 mg/mL of USP Clidinium Bromide RS in methanol. Sonicate to dissolve.
Standard solution: 0.01 mg/mL of USP Chlordiazepoxide Hydrochloride RS and 0.005 mg/mL of USP Clidinium Bromide RS from the Standard stock solution, in Medium. Store at 5°.
Sample solution: Pass a portion of the solution under test through a suitable PVDF filter of 0.45-μm pore size, discarding the first 5 mL of filtrate. Store at 5°.
Chromatographic system
(See Chromatography 〈621〉, System Suitability.) Mode: LC
Detector: UV 212 nm
Column: 4.6-mm × 15-cm; 4.0-μm packing L1 Temperatures
Autosampler: 5°
Column: 50° Flow rate: 1 mL/min
Injection volume: 25 µL
Run time: NLT 2 times the retention time of chlordiazepoxide System suitability
Sample: Standard solution
[NOTE—The relative retention times for clidinium and chlordiazepoxide are about 0.43 and 1.0, respectively.] Suitability requirements
Resolution: NLT 5.0 between clidinium and chlordiazepoxide Tailing factor: NMT 2.0 for clidinium and chlordiazepoxide
Relative standard deviation: NMT 2.0% for clidinium and chlordiazepoxide
Analysis
Samples: Standard solution and Sample solution
Calculate the percentage of the labeled amount of chlordiazepoxide hydrochloride (C H ClN O · HCl) and clidinium bromide (C H BrNO ) dissolved:
Result = (r /r ) × C × V × (1/L) × 100
= peak response of chlordiazepoxide or clidinium from the Sample solution
= peak response of chlordiazepoxide or clidinium from the Standard solution
= concentration of USP Chlordiazepoxide Hy drochloride RS or USP Clidinium Bromide RS in the Standard solution (mg/mL)
V = volume of Medium, 500 mL
L = label claim for chlordiazepoxide hydrochloride or clidinium bromide (mg/Capsule)
Tolerances: NLT 80% (Q ) each of the labeled amount of chlordiazepoxide hydrochloride (C H ClN O · HCl) and clidinium bromide (C22H26BrNO3) is dissolved.▲ (RB 5-Nov-2021)
UNIFORMITY OF DOSAGE UNITS 〈905〉: Meet the requirements
5 IMPURITIES
5.1 LIMIT OF CHLORDIAZEPOXIDE RELATED COMPOUND A AND 2-AMINO-5-CHLOROBENZOPHENONE
Standard solution A: 1 mg/mL of USP Chlordiazepoxide Related Compound A RS in acetone Standard solution B: 50 µg/mL of USP 2-Amino-5-chlorobenzophenone RS in acetone
Sample solution: Transfer an amount equivalent to 25 mg of chlordiazepoxide hydrochloride from Capsule contents to a 10-mL conical flask,
add 2.5 mL of acetone, and shake. Allow any undissolved particles to settle, and use the supernatant.
5.1.1 Chromatographic system
(See Chromatography 〈621〉, General Procedures, Thin-Layer Chromatography.) Adsorbent: 0.25-mm layer of chromatographic silica gel
Application volume: 50 µL for the Sample solution, 15 µL for Standard solution A, and 10 µL for Standard solution B Developing solvent system: Ethyl acetate
Spray reagent: 2 N sulfuric acid Analysis
Samples: Standard solutions and Sample solution
Proceed as directed in the chapter. Develop the chromatogram in a chromatographic chamber (not previously saturated with the developing solvent) in the Developing solvent system until the solvent front has moved three-fourths of the length of the plate. Remove the plate from the developing chamber, mark the solvent front, and allow the solvent to evaporate. Locate the spots on the plate by lightly spraying with Spray reagent. Dry at 105° for 15 min, and then spray in succession with sodium nitrite solution (1 in 1000), ammonium sulfamate solution (1 in 200), and N-(1-naphthyl)ethylenediamine dihydrochloride solution (1 in 1000).
Acceptance criteria: Any spots from the Sample solution are not greater in size or intensity than the spots at the respective R values
produced by the Standard solutions, corresponding to NMT 3.0% of chlordiazepoxide related compound A and to NMT 0.1% of 2-amino-5- chlorobenzophenone.
5.2 LIMIT OF CLIDINIUM BROMIDE RELATED COMPOUND A
Extracting solvent mixture: Dehydrated alcohol and cyclohexane (1:1)
Identification solution: Dissolve 50 mg of USP Clidinium Bromide RS in 1 mL of 0.1 N methanolic hydrochloric acid. To this solution add 20 µL of a solution of 25 mg/mL of USP Clidinium Bromide Related Compound A RS in methanol. Prepare this solution at the time of use.
Standard solution: 50 mg/mL of USP Clidinium Bromide RS in 0.1 N methanolic hydrochloric acid. [NOTE—Prepare this solution at the time of use.]
Sample solution: Empty a number of Capsules, equivalent to 25 mg of clidinium bromide, into a glass-stoppered centrifuge tube, and add 5 mL of the Extracting solvent mixture. Heat the tube gently, with shaking, to 50°, centrifuge, and decant the clear supernatant into a second tube. Repeat the addition of Extracting solvent mixture twice, heating, centrifuging, and decanting as before, and combine the three extracts in a single tube. Gently heating, evaporate the combined extracts under a stream of nitrogen to dryness. Dissolve the residue in 0.5 mL of methanol.
5.2.1 Chromatographic system
(See Chromatography 〈621〉, General Procedures, Thin-Layer Chromatography.) Adsorbent: 0.25-mm layer of chromatographic silica gel mixture Application volume: 20 µL
Developing solvent system: Acetone, methanol, water, and hy drochloric acid (70:20:5:5)
Spray reagent: Dissolve 850 mg of Bismuth subnitrate in a mixture of 10 mL of glacial acetic acid and 40 mL of water. In a separate container, dissolve 20 g of potassium iodide in 50 mL of water. Mix the two solutions, and dilute with dilute sulfuric acid (1 in 10) to 500 mL. Add 7.5 ± 2.5 g of iodine, and mix until solution is complete.
Chromatographic plates: Predevelop suitable thin-layer chromatographic plates by placing in a chromatographic chamber saturated with Developing solvent system, and allow the Developing solvent system to move 15 cm. Remove the plates from the chamber, dry at 105° for 15 min, and cool.
5.2.2 Analysis
Samples: Identification solution, Standard solution, and Sample solution
Proceed as directed in the chapter. Place the plates in an unsaturated chromatographic chamber containing freshly prepared Developing solvent system, and develop the chromatogram until the solvent front has moved 15 cm. Remove the plates, and dry at 105° for 10 min. Cool to room temperature, and spray with Spray reagent. Any spot in the chromatogram of the Sample solution occurring at an R value of 0.4 is not greater in size or intensity than the corresponding spot in the chromatogram of the Identification solution; and the Standard solution shows no spot at the R value corresponding to that of clidinium bromide related compound A.
Acceptance criteria: NMT 1.0% of clidinium bromide related compound A
6 ADDITIONAL REQUIREMENTS
PACKAGING AND STORAGE: Preserve in tight, light-resistant containers. Add the following:
LABELING: When more than one Dissolution test is used, the labeling states the Dissolution test used only if Test 1 is not used.▲ (RB 5-Nov-2021)
USP REFERENCE STANDARDS 〈11〉
USP 2-Amino-5-chlorobenzophenone RS
USP Chlordiazepoxide Hydrochloride RS
USP Chlordiazepoxide Related Compound A RS
USP Clidinium Bromide RS
USP Clidinium Bromide Related Compound A RS
