Midazolam Injection
DEFINITION
Midazolam Injection is a sterile solution of Midazolam Hydrochloride in Water for Injection or of Midazolam in Water for Injection prepared with the aid of Hydrochloric Acid. It contains the equivalent of NLT 90.0% and NMT 110.0% of the labeled amount of midazolam (C18H13ClFN3). It may contain Sodium Chloride, Benzyl Alcohol, and/or a chelating agent.
IDENTIFICATION
• The retention time of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay.
ASSAY
[Note—Protect all prepared Standard and sample solutions from light. ]
• Procedure
Buffer:
6.7 g/L of dibasic sodium phosphate heptahydrate in water. Adjust with phosphoric acid to a pH of 5.0 ± 0.1.
Solution A:
Prepare a filtered and degassed mixture of acetonitrile, methanol and Buffer (8:3:9).
Solution B:
Acetonitrile and Buffer (3:1)
Mobile phase:
See the gradient table below.
| Time (min) |
Solution A (%) |
Solution B (%) |
|---|---|---|
| 0 | 100 | 0 |
| 15 | 100 | 0 |
| 20 | 0 | 100 |
| 35 | 0 | 100 |
| 37 | 100 | 0 |
| 45 | 100 | 0 |
Standard solution:
Dissolve USP Midazolam RS in about 2 mL of methanol, and dilute quantitatively, and stepwise if necessary, with Solution A to obtain a 0.2-mg/mL solution.
Sample solution:
[Note—The midazolam present in the Injection converts from the open-ring form to the closed-ring form when diluted with Solution A. The midazolam potency is determined based on the peak area of the closed-ring form. It takes approximately 60 min at 40
or 2–3 h at room temperature to complete the conversion. The Standard solution is not subject to this conversion process. ] Transfer a volume of Injection to a suitable volumetric flask, and dilute with Solution A to obtain a solution containing about 0.2 mg/mL of midazolam. Transfer the resulting solution into suitable crimp top vials, seal tightly, and heat at about 40 for 60 min. Allow this solution to cool to room temperature before injection.
or 2–3 h at room temperature to complete the conversion. The Standard solution is not subject to this conversion process. ] Transfer a volume of Injection to a suitable volumetric flask, and dilute with Solution A to obtain a solution containing about 0.2 mg/mL of midazolam. Transfer the resulting solution into suitable crimp top vials, seal tightly, and heat at about 40 for 60 min. Allow this solution to cool to room temperature before injection.
Chromatographic system
Mode:
LC
Detector:
UV 254 nm
Column:
4.6-mm × 25-cm; packing L1
Flow rate:
1.0 mL/min
Injection size:
50 µL
System suitability
Sample:
Standard solution
Suitability requirements
Column efficiency:
NLT 5500 theoretical plates
Tailing factor:
NMT 2.5
Relative standard deviation:
NMT 2.0%
Analysis
Samples:
Standard solution and Sample solution
Calculate the percentage of labeled amount of C18H13ClFN3 in the portion of Injection 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 | = | = nominal concentration of Midazolam in the Sample solution (mg/mL) |
Acceptance criteria:
90.0%–110.0%
IMPURITIES
Organic Impurities
[Note—Protect all prepared Standard and sample solutions from light. ]
• Procedure
Buffer, Solution A, Solution B, Mobile phase, Sample solution, and Chromatographic system:
Proceed as directed in the Assay.
Standard stock solution:
Use Standard solution in the Assay.
Standard solution:
0.5 µg/mL USP Midalozam RS in Solution A from Standard stock solution
Control solution:
0.1 µg/mL USP Midalozam RS in Solution A from Standard solution
System suitability
Samples:
Standard solution and Control solution
Suitability requirements
Tailing factor:
NMT 2.5 for midalozam peak, Standard solution
Column efficiency:
NLT 5500 theoretical plates, Standard solution
Signal-to-noise ratio:
NLT 10, Control solution
Relative standard deviation:
NMT 8.0%, Standard solution
Analysis
Samples:
Standard solution and Sample solution
Calculate the percentage of each impurity in the portion of Injection taken:
Result = (rU/rS) × (CS/CU) × (1/F) × 100
| rU | = | = peak response of the individual impurity from the Sample solution |
| rS | = | = peak response of midalozam from the Standard solution |
| CS | = | = concentration of USP Midalozam RS in the Standard solution (mg/mL) |
| CU | = | = nominal concentration of Midalozam in the Sample solution (mg/mL) |
| F | = | = relative response factor; 0.51 for the peak eluting at a relative retention between 0.79 and 0.97 with respect to midazolam; 1.0 for all other peaks |
Acceptance criteria
Individual known impurity:
NMT 0.5%
Individual unknown impurity:
NMT 0.1%
Total impurities:
NMT 1.0%
[Note—Disregard all solvent- and excipient-related peaks. ]
SPECIFIC TESTS
• Benzyl Alcohol Content (if present)
Buffer:
3.4 g/L of monobasic sodium phosphate in water. Adjust with phosphoric acid to a pH of 3.5.
Mobile phase:
Acetonitrile and Buffer (7:13)
System suitability solution:
0.05 mg/mL of USP Midazolam RS and 0.5 mg/mL of USP Benzyl Alcohol RS in Mobile phase
Standard solution:
0.5 mg/mL of USP Benzyl Alcohol RS in Mobile phase
Sample solution:
Transfer a measured volume of Injection to a suitable volumetric flask. Dilute with Mobile phase to obtain a concentration of about 0.5 mg/mL of benzyl alcohol, based on the labeled content of benzyl alcohol in the Injection.
Chromatographic system
Mode:
LC
Detector:
UV 254 nm
Column:
4.6-mm × 25-cm; L1 packing
Flow rate:
1.0 mL/min
Injection size:
50 µL
System suitability
Sample:
System suitability solution
Suitability requirements
Resolution:
NLT 6.0 between benzyl alcohol and midazolam
Tailing factor:
NMT 2.0 for benzyl alcohol
Relative standard deviation:
NMT 2.0% for benzyl alcohol
Analysis
Samples:
Standard solution and Sample solution
Calculate the percentage of the labeled amount of benzyl alcohol in the volume of Injection taken:
Result = (rU/rS) × (CS/CU) × 100
| rU | = | = peak response of benzyl alcohol from the Sample solution |
| rS | = | = peak response of benzyl alcohol from the Standard solution |
| CS | = | = concentration of USP Benzyl Alcohol RS in the Standard solution (mg/mL) |
| CU | = | = nominal concentration of benzyl alcohol in the Sample solution (mg/mL) |
Acceptance criteria:
The content of benzyl alcohol meets the requirements for Added Substances under Injections 
1
.
1.
• Particulate Matter in Injections 788:
Meets the requirements for small-volume injections
788:
Meets the requirements for small-volume injections
• Bacterial Endotoxins Test 85:
It contains NMT 8.33 USP Endotoxin Units/mg of midazolam.
85:
It contains NMT 8.33 USP Endotoxin Units/mg of midazolam.
• pH 791:
2.5–3.7
791:
2.5–3.7
• Sterility Tests 71:
Meets the requirements
71:
Meets the requirements
• Other Requirements:
It meets the requirements for Injections 1.
1.
ADDITIONAL REQUIREMENTS
• Packaging and Storage:
Preserve in single-dose containers, preferably of Type 1 glass. Store between 15 and 30.
and 30.
• Labeling:
Label to indicate the vehicle used and the names and concentrations of any added preservatives. Indicate if the product is preservative free.
') + '&img=../../images/v35300/cas-100-51-6.gif',600,500);)
Auxiliary Information—
Please check for your question in the FAQs before contacting USP.
| Topic/Question | Contact | Expert Committee |
|---|---|---|
| Monograph | Mary S. Waddell
Scientific Liaison 1-301-816-8124 |
(SM42010) Monographs - Small Molecules 4 |
| 85 |
Radhakrishna S Tirumalai, Ph.D.
Principal Scientific Liaison 1-301-816-8339 |
(GCM2010) General Chapters - Microbiology |
| 71 |
Radhakrishna S Tirumalai, Ph.D.
Principal Scientific Liaison 1-301-816-8339 |
(GCM2010) General Chapters - Microbiology |
| Reference Standards | RS Technical Services 1-301-816-8129 rstech@usp.org |
USP35–NF30 Page 3920
Pharmacopeial Forum: Volume No. 34(3) Page 635