Midazolam

This article is compiled based on the United States Pharmacopeia (USP) – 2025 Edition

Issued and maintained by the United States Pharmacopeial Convention (USP)

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C₁₈H₁₃ClFN₃ 325.77

4-H-Imidazo[1,5-a][1,4]benzodiazepine, 8-chloro-6-(2-fluorophenyl)-1-methyl;

8-Chloro-6-(o-fluorophenyl)-1-methyl-4H-imidazo[1,5-a][1,4]benzodiazepine  CAS RN®: 59467-70-8; UNII: R60L0SM5BC.

1 DEFINITION

Midazolam contains NLT 98.5% and NMT 101.5% of C₁₈H₁₃ClFN₃, calculated on the dried basis.

2 IDENTIFICATION

Change to read:

A. Spectroscopic Identification Tests 〈197〉, Infrared Spectroscopy: 197K

B. The retention time of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay.

3 ASSAY

3.1 Procedure

Buffer: 7.7 g/L of ammonium acetate in water. Adjust with glacial acetic acid to a pH of 5.5 ± 0.1.

Mobile phase: Acetonitrile and Buffer (1:2)

Standard solution: 0.04 mg/mL of USP Midazolam RS in Mobile phase

Sample solution: 0.04 mg/mL of Midazolam in Mobile phase

Chromatographic system

• (See Chromatography 〈621〉, System Suitability.)
• Mode: LC
• Detector: UV 254 nm
• Column: 4.6-mm × 25-cm; 5-µm packing L60
• Flow rate: 1.5 mL/min
• Injection size: 25 µL

System suitability

• Sample: Standard solution
• Suitability requirements
• Column efficiency: NLT 10,000 theoretical plates
• Tailing factor: NMT 2.0
• Relative standard deviation: NMT 2.0%

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of C₁₈H₁₃ClFN₃ in the portion of Midazolam taken:

Result = (rᵤ/rₛ) × (Cₛ/Cᵤ) × 100

rᵤ = peak response from the Sample solution

rₛ = peak response from the Standard solution

Cₛ = concentration of USP Midazolam RS in the Standard solution (mg/mL)

Cᵤ = concentration of Midazolam in the Sample solution (mg/mL)

Acceptance criteria: 98.5%–101.5% on the dried basis

4 IMPURITIES

4.1 Inorganic Impurities

Residue on Ignition 〈281〉: NMT 0.1%

4.2 Organic Impurities

4.2.1 Procedure

Buffer, Mobile phase, Standard solution, and Chromatographic system: Proceed as directed in the Assay.

Sensitivity check solution: Dilute the Standard solution with Mobile phase to obtain a 0.2-µg/mL solution.

Sample solution: 0.2 mg/mL of Midazolam in Mobile phase

System suitability

Samples: Standard solution and Sensitivity check solution

Suitability requirements

• Column efficiency: NLT 10,000 theoretical plates, Standard solution
• Tailing factor: NMT 2.0, Standard solution
• Relative standard deviation: NMT 2.0%, Standard solution
• Peak ratio: The ratio of the area of the midazolam peak of the Standard solution to the area of the midazolam peak of the Sensitivity check solution should be within 160–240.

Analysis

Sample: Sample solution

Calculate the percentage of each impurity in the portion of Midazolam taken:

Result = (rᵤ/F)/[Σ(rᵤ/F) + r_T] × 100

rᵤ = peak response of each individual impurity from the Sample solution

r_T = peak response of Midazolam from the Sample solution

F = relative response factor (see Impurity Table 1)

Acceptance criteria: See Impurity Table 1.

Impurity Table 1

ᵃ 8-Chloro-3a,4-dihydro-6-(2-fluorophenyl)-1-methyl-3H-imidazo[1,5-a][1,4]-benzodiazepine.

ᵇ 8-Chloro-6-(2-fluorophenyl)-3a,4,5,6-tetrahydro-1-methyl-3H-imidazo[1,5-a][1,4]-benzodiazepine.

ᶜ 2-Aminomethyl-7-chloro-2,3-dihydro-5-(2-fluorophenyl)-1H-1,4-benzodiazepine.

ᵈ 8-Chloro-6-(2-fluorophenyl)-1-methyl-4H-imidazo[1,5-a][1,4]-benzodiazepine-5-oxide.

ᵉ 7-Chloro-1,3-dihydro-2-nitromethylene-5-(2-fluorophenyl)-2H-1,4-benzodiazepine-4-oxide.

ᶠ 8-Chloro-6-(2-fluorophenyl)-5,6-dihydro-1-methyl-4H-imidazo[1,5-a][1,4]-benzodiazepine.

ᵍ 8-Chloro-6-phenyl-1-methyl-4H-imidazo-[1,5-a][1,4]-benzodiazepine.

ʰ 8-Chloro-6-(2-fluorophenyl)-1-methyl-6H-imidazo[1,5-a][1,4]-benzodiazepine.

5 SPECIFIC TESTS

Loss on Drying 〈731〉: Dry a sample at 105° for 2 h: it loses NMT 0.5% of its weight.

6 ADDITIONAL REQUIREMENTS

Packaging and Storage: Preserve in tight, light-resistant containers.

USP Reference Standards 〈11〉

USP Midazolam RS